CN104557568A - Nicotinic receptor non-competitive antagonists - Google Patents

Nicotinic receptor non-competitive antagonists Download PDF

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CN104557568A
CN104557568A CN201510013360.XA CN201510013360A CN104557568A CN 104557568 A CN104557568 A CN 104557568A CN 201510013360 A CN201510013360 A CN 201510013360A CN 104557568 A CN104557568 A CN 104557568A
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compound
alkyl
disease
illness
rings
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CN104557568B (en
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S·R·阿基雷迪
B·S·巴蒂
R·J·海姆斯特拉
M·S·梅尔文
J·斯皮克
Y·肖
D·约翰内斯
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Catalyst Biosciences Inc
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Targacept Inc
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    • C07C211/33—Compounds containing amino groups bound to a carbon skeleton having amino groups bound to carbon atoms of rings other than six-membered aromatic rings
    • C07C211/39—Compounds containing amino groups bound to a carbon skeleton having amino groups bound to carbon atoms of rings other than six-membered aromatic rings of an unsaturated carbon skeleton
    • C07C211/41—Compounds containing amino groups bound to a carbon skeleton having amino groups bound to carbon atoms of rings other than six-membered aromatic rings of an unsaturated carbon skeleton containing condensed ring systems
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    • A61K31/00—Medicinal preparations containing organic active ingredients
    • A61K31/33—Heterocyclic compounds
    • A61K31/395—Heterocyclic compounds having nitrogen as a ring hetero atom, e.g. guanethidine or rifamycins
    • A61K31/397—Heterocyclic compounds having nitrogen as a ring hetero atom, e.g. guanethidine or rifamycins having four-membered rings, e.g. azetidine
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
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    • A61K31/33—Heterocyclic compounds
    • A61K31/395—Heterocyclic compounds having nitrogen as a ring hetero atom, e.g. guanethidine or rifamycins
    • A61K31/40—Heterocyclic compounds having nitrogen as a ring hetero atom, e.g. guanethidine or rifamycins having five-membered rings with one nitrogen as the only ring hetero atom, e.g. sulpiride, succinimide, tolmetin, buflomedil
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    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P1/00—Drugs for disorders of the alimentary tract or the digestive system
    • A—HUMAN NECESSITIES
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    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
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    • A61P13/10—Drugs for disorders of the urinary system of the bladder
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P25/00—Drugs for disorders of the nervous system
    • A61P25/24—Antidepressants
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P25/00—Drugs for disorders of the nervous system
    • A61P25/26—Psychostimulants, e.g. nicotine, cocaine
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P25/00—Drugs for disorders of the nervous system
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    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P25/00—Drugs for disorders of the nervous system
    • A61P25/30—Drugs for disorders of the nervous system for treating abuse or dependence
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    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
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    • C07—ORGANIC CHEMISTRY
    • C07C—ACYCLIC OR CARBOCYCLIC COMPOUNDS
    • C07C211/00—Compounds containing amino groups bound to a carbon skeleton
    • C07C211/33—Compounds containing amino groups bound to a carbon skeleton having amino groups bound to carbon atoms of rings other than six-membered aromatic rings
    • C07C211/34—Compounds containing amino groups bound to a carbon skeleton having amino groups bound to carbon atoms of rings other than six-membered aromatic rings of a saturated carbon skeleton
    • C07C211/38—Compounds containing amino groups bound to a carbon skeleton having amino groups bound to carbon atoms of rings other than six-membered aromatic rings of a saturated carbon skeleton containing condensed ring systems
    • C—CHEMISTRY; METALLURGY
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    • C07D—HETEROCYCLIC COMPOUNDS
    • C07D205/00—Heterocyclic compounds containing four-membered rings with one nitrogen atom as the only ring hetero atom
    • C07D205/02—Heterocyclic compounds containing four-membered rings with one nitrogen atom as the only ring hetero atom not condensed with other rings
    • C07D205/04—Heterocyclic compounds containing four-membered rings with one nitrogen atom as the only ring hetero atom not condensed with other rings having no double bonds between ring members or between ring members and non-ring members
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    • C07—ORGANIC CHEMISTRY
    • C07D—HETEROCYCLIC COMPOUNDS
    • C07D207/00—Heterocyclic compounds containing five-membered rings not condensed with other rings, with one nitrogen atom as the only ring hetero atom
    • C07D207/02—Heterocyclic compounds containing five-membered rings not condensed with other rings, with one nitrogen atom as the only ring hetero atom with only hydrogen or carbon atoms directly attached to the ring nitrogen atom
    • C07D207/04—Heterocyclic compounds containing five-membered rings not condensed with other rings, with one nitrogen atom as the only ring hetero atom with only hydrogen or carbon atoms directly attached to the ring nitrogen atom having no double bonds between ring members or between ring members and non-ring members
    • C07D207/06—Heterocyclic compounds containing five-membered rings not condensed with other rings, with one nitrogen atom as the only ring hetero atom with only hydrogen or carbon atoms directly attached to the ring nitrogen atom having no double bonds between ring members or between ring members and non-ring members with radicals, containing only hydrogen and carbon atoms, attached to ring carbon atoms
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    • C07D295/00—Heterocyclic compounds containing polymethylene-imine rings with at least five ring members, 3-azabicyclo [3.2.2] nonane, piperazine, morpholine or thiomorpholine rings, having only hydrogen atoms directly attached to the ring carbon atoms
    • C07D295/02—Heterocyclic compounds containing polymethylene-imine rings with at least five ring members, 3-azabicyclo [3.2.2] nonane, piperazine, morpholine or thiomorpholine rings, having only hydrogen atoms directly attached to the ring carbon atoms containing only hydrogen and carbon atoms in addition to the ring hetero elements
    • C07D295/027—Heterocyclic compounds containing polymethylene-imine rings with at least five ring members, 3-azabicyclo [3.2.2] nonane, piperazine, morpholine or thiomorpholine rings, having only hydrogen atoms directly attached to the ring carbon atoms containing only hydrogen and carbon atoms in addition to the ring hetero elements containing only one hetero ring
    • C07D295/033—Heterocyclic compounds containing polymethylene-imine rings with at least five ring members, 3-azabicyclo [3.2.2] nonane, piperazine, morpholine or thiomorpholine rings, having only hydrogen atoms directly attached to the ring carbon atoms containing only hydrogen and carbon atoms in addition to the ring hetero elements containing only one hetero ring with the ring nitrogen atoms directly attached to carbocyclic rings

Abstract

The present invention relates to compounds or medically-acceptable salt that modulate nicotinic receptors as non-competitive antagonists, methods for their synthesis, methods for use, and their pharmaceutical compositions, wherein R1 and R2 respectively and individually are H,C1-6 alkyl group, or aryl group-replaced C1-6 alkyl group, or R1 and R2 and nitrogen atoms connected to R1 and R2 form 3 to 8 rings, and the rings can be replaced by any of the below items: C1-6 alkyl group, aryl group, C1-6 alkoxy group, or aryloxy substituent, and dotted lines refer to optional double bonds.

Description

NAChR noncompetitive antaganist
The divisional application of the application's to be female case be Chinese invention patent application 201180026122.2.
Invention field
The present invention relates to the compound regulating nAChR as noncompetitive conditioning agent (such as, noncompetitive antaganist), their synthetic method, using method, and their pharmaceutical composition.
Background of invention
NAChR be a large amount of dystopy regulate the external source of its function and the target of endogenous compounds.See Arias, H.R., Binding sites for exogenous and endogenousnon-competitive inhibitors of the nicotinic acetylcholine receptor, Biochimica et Biophysica Acta-Reviews on Biomembranes 1376:173-220 (1998) and Arias, H.R., Bhumireddy, P., Anesthetics as chemicaltools to study the structure and function of nicotinic acetylcholinereceptors, Current Protein & Peptide Science 6:451-472 (2005). the function of nAChR can by being called that the compound that noncompetitive conditioning agent comprises the configurations of noncompetitive antaganist is minimized or blocks (see summary Arias, H.R., Bhumireddy, P., Bouzat, C., Molecular mechanisms and binding site locations for noncompetitiveantagonists of nicotinic acetylcholine receptors.The InternationalJournal of Biochemistry & Cell Biology 38:1254-1276 (2006)).
Noncompetitive conditioning agent comprises the compound of the configurations of wide region, and it suppresses function of receptors by working in the one or more sites being different from ortho position resistance (orthosteric) binding site.Regulation is verified is high complexity.The mechanism of noncompetitive modulator effect and binding affinity is different (people such as Arias, 2006) to each nicotinic receptor subtypes.Noncompetitive conditioning agent can be worked by least two kinds of different mechanism: allosteric mechanism and/or steric hindrance mechanism.
Allosteric antagonist mechanism involves the combination of noncompetitive antaganist and acceptor and non-stabilization of carrying out conformation state that is tranquillization or desensitized state, and/or the increase of receptor desensitization speed.
In contrast, the direct representative of steric hindrance mechanism is the antagonist molecules physically blocking ionic channel.Described antagonist can be called noncompetitive channel modulators (NCMs).Some suppresses acceptor like this: combine in hole when acceptor is in open state, physically block iontophoretic injection thus.Some only works as pure open channel blocker, and other block passage that is open and that close.The mechanism suppression ionic flux that described antagonist combines by not being involved in resistance site, ortho position.
Show, group of barbiturates, the narcotic that dissociates, thymoleptic and some steroid suppress nAChR by comprising open and that closed channel is closed allosteric mechanism.The model such to the research support of group of barbiturates, wherein combines and occurs in the opening of acceptor and closed state, causes closing of ionic current.See Dilger, J.P., Boguslavsky, R., Barann, M., Katz, T., Vidal, A.M., Mechanisms of barbiturate inhibition of acetylcholine receptorchannels, Journal General Physiology 109:401-414 (1997).Although the inhibition effect of local anesthetic to nerve conduction is mainly mediated by closed voltage-gated sodium channels, nAChR is also the target of local anesthetic.See Arias, H.R., Role of localanesthetics on both cholinergic and serotonergic ionotropic receptors, Neuroscience and Biobehavioral Reviews 23:817-843 (1999) and Arias, H.R. & Blanton, M.P., Molecular and physicochemical aspects of localanesthetics acting on nicotinic acetylcholine receptor-containingmembranes, Mini Reviews in Medicinal Chemistry 2:385-410 (2002).
Such as, tetracaine is preferentially bonded to receptor channel in tranquillization state.The narcotic that dissociates suppresses several neuronic-type nAChRs within the scope of clinical concentration, example is such as phencyclidine (PCP) (Connolly, J., Boulter, J., & Heinemann, S.F., Alpha 4-beta 2and othernicotinic acetylcholine receptor subtypes as targets of psychoactive andaddictive drugs, British Journal of Pharmacology 105:657-666 (1992)), ketamine (Flood, P. & Krasowski M.D., Intravenous anestheticsdifferentially modulate ligand-gated ion channels, Anesthesiology 92:1418-1425 (2000), and Ho, K.K. & Flood, P., Single amino acid residue inthe extracellular portion of transmembrane segment 2in the nicotinic α 7acetylcholine receptor modulates sensitivity to ketamine, Anesthesiology100:657-662 (2004)), with Dizocilpine (Krasowski, M.D., & Harrison, N.L., General anaesthetic actions on ligand-gated ion channels, Cellular andMolecular Life Sciences 55:1278-1303 (1999)).Research is pointed out, the narcotic that dissociates is bonded to the single or overlapping place in tranquillization ionic channel, and to show that in receptor channel ketamine/PCP position partly overlaps tetracaine binding site.Dizocilpine, also referred to as MK-801, be dissociate narcotic and anticonvulsive drug, it also acts on different nAChRs as noncompetitive antaganist.Dizocilpine it was reported it is the open channel blocker of α 4 β 2 neuronal nicotinic receptor.See Buisson, B., & Bertrand, D., Open-channel blockers at the human α 4 β 2neuronal nicotinic acetylcholine receptor, Molecular Pharmacology 53:555-563 (1998).
Except they know to except the effect of monoamine and serotonin reuptake system, thymoleptic are vision-control nAChR also.Previous research display tricyclics serves as noncompetitive antaganist.See Gumilar, F., Arias, H.R., Spitzmaul, G., Bouzat, C., Molecular mechanisms of inhibition of nicotinic acetylcholine receptorsby tricyclic antidepressants.Neuropharmacology 45:964-76 (2003). fluoxetine is reported as selective serotonin reuptake inhibitor (SSRI) by the membrane current increasing desensitization speed and/or suppress activating muscle or neuronic nAChR to cause by induction channel enclosed with non-competitive fashion Deng people.See j., Awad, J.N., & Miledi, R., Blockage of muscle and neuronal nicotinicacetylcholine receptors by fluoxetine (Prozac), Proceedings of theNational Academy of Sciences USA 94:2041-2044 (1997); With j., Vazquez-Gomez, E., & Miledi, R., Combined actionsof zinc and fluoxetine on nicotinic acetylcholine receptors, ThePharmacogenomics Journal 4:388-393 (2004).It is classical noncompetitive nicotinic receptor antagonist that approval is in advance used for the treatment of hypertensive mecamylamine, and also knows by blocking ionic channel suppression function of receptors.See Giniatullin, R.A., Sokolova, E.M., DiAngelantonio, S., Skorinkin, A., Talantova, M.V., Nistri, A.Rapid Reliefof Block by Mecamylamine of Neuronal Nicotinic AcetylcholineReceptors of Rat Chromaffin Cells In Vitro:An Electrophysiological andModeling Study.Molecular Pharmacology 58:778-787 (2000).
Brief summary of the invention
The present invention includes formula I:
Formula I
Wherein
R 1and R 2h, C individually separately 1-6alkyl, or the C of aryl-replacement 1-6alkyl, or R 1and R 2the nitrogen-atoms be connected to them is combined to form 3 to 8 rings, and this ring optionally can use following replacement: C 1-6alkyl, aryl, C 1-6alkoxyl group, or aryloxy substituting group;
R 3h, C 1-6alkyl, or C 1-6the C of alkoxyl group-replacement 1-6alkyl;
R 4, R 5, R 6and R 7h, C individually separately 1-6alkyl, or C 1-6alkoxyl group;
L 1be selected from following linker kind: CR 8r 9, CR 8r 9cR 10r 11, and O;
L 2be selected from following linker kind: CH 2, CH 2cH 2, CH 2cH 2cH 2, or CH 2cH 2cH 2cH 2;
R 8, R 9, R 10and R 11hydrogen or C individually separately 1-6alkyl; With
Dotted line points out optional double bond;
Or its pharmacy acceptable salt.
The present invention includes pharmaceutical composition, it comprises the compounds of this invention or its pharmacy acceptable salt.Pharmaceutical composition of the present invention can be used in treatment or prevents illness miscellaneous or obstacle, particularly those obstacles of characterizing of nicotine cholinergic drug neurotransmission dysfunction or nicotine cholinergic drug deterioration of neurons.
The present invention includes for needing mammalian therapeutic or the prevention obstacle and dysfunction such as CNS obstacle and dysfunction for the treatment of, and be used for the treatment of or prevent that some illness such as eases the pain, the method for hypertension and inflammation.Described method involves and gives to treat the compounds of this invention of significant quantity to experimenter and comprise its salt, or comprises the pharmaceutical composition of described compound.
Detailed Description Of The Invention
I. compound
One embodiment of the present invention comprise formula I:
Formula I
Wherein
R 1and R 2h, C individually separately 1-6alkyl, or the C of aryl-replacement 1-6alkyl, or R 1and R 2the nitrogen-atoms be connected to them is combined to form 3 to 8 rings, and this ring optionally can use following replacement: C 1-6alkyl, aryl, C 1-6alkoxyl group, or aryloxy substituting group;
R 3h, C 1-6alkyl, or C 1-6the C of alkoxyl group-replacement 1-6alkyl;
R 4, R 5, R 6and R 7h, C individually separately 1-6alkyl, or C 1-6alkoxyl group;
L 1be selected from following linker kind: CR 8r 9, CR 8r 9cR 10r 11, and O;
L 2be selected from following linker kind: CH 2, CH 2cH 2, CH 2cH 2cH 2, or CH 2cH 2cH 2cH 2;
R 8, R 9, R 10and R 11hydrogen or C individually separately 1-6alkyl; With
Dotted line points out optional double bond;
Or its pharmacy acceptable salt.
In one embodiment, R 1be H and R 2c 1-6alkyl.In one embodiment, R 3c 1-6alkyl.In one embodiment, R 4, R 5, R 6and R 7each H naturally.In one embodiment, L 1cR 8r 9, and R 8and R 9each hydrogen naturally.In one embodiment, L 2cH 2cH 2.In one embodiment, dotted line is singly-bound.
An aspect of of the present present invention comprises pharmaceutical composition, and it comprises the compounds of this invention and pharmaceutically acceptable carrier.
An aspect of of the present present invention comprises, particularly by using noncompetitive conditioning agent (such as non-competing antagonist) to include but not limited to channel blocker, be used for the treatment of or prevent the method for disease or the illness mediated by neuronic nAChR, comprise administration the compounds of this invention.In one embodiment, described disease or illness are CNS obstacles.In yet, described disease or illness are inflammation or Inflammatory response.In yet, described disease or illness are pain.In yet, described disease or illness are neovascularization.In yet, described disease or illness are hypertension.In yet, described disease or illness are other obstacles described herein.
An aspect of of the present present invention comprises the purposes of the compounds of this invention for the preparation of medicine, and described medicine is particularly by using noncompetitive antaganist such as channel blocker to be used for the treatment of or preventing the disease that mediated by neuronic nAChR or illness.In one embodiment, described disease or illness are CNS obstacles.In yet, described disease or illness are inflammation or Inflammatory response.In yet, described disease or illness are pain.In yet, described disease or illness are neovascularization.In yet, described disease or illness are hypertension.In yet, described disease or illness are other obstacles described herein.
An aspect of of the present present invention comprises the compounds of this invention as active therapeutic agent.Thus, comprise on the one hand particularly by using noncompetitive antaganist such as channel blocker be used for the treatment of or prevent the compounds of this invention of disease or the illness mediated by neuronic nAChR.In one embodiment, described disease or illness are CNS obstacles.In yet, described disease or illness are inflammation or Inflammatory response.In yet, described disease or illness are pain.In yet, described disease or illness are neovascularization.In yet, described disease or illness are hypertension.In yet, described disease or illness are other obstacles described herein.
Special disease or illness comprise depression, comprise major depressive disorder, hypertension, irritable bowel syndrome (IBS), comprise IBS-D (diarrhoea is preponderated), bladder hyperactivity (OAB), and habituation, comprise smoking and stop.
Scope of the present invention comprises whole combinations of each side and each embodiment.
Following definitions is intended to clarification but is not limited to defined term.If special term used herein does not define especially, then described term should not thought uncertain.On the contrary, term uses with its implication accepted.In this specification sheets whole, preferred atomic ratio represents as the quantity of carbon atom: such as phrase " C x-yalkyl ", it refers to the alkyl defined herein containing specifying carbonatoms.Similar nomenclature is also applicable to other preferred term and scope.Thus, such as, C 1-6alkyl representative is containing the straight chain of 1 to 6 carbon atom or the chain hydrocarbon of branching.
Term as used herein " alkyl " refers to the chain hydrocarbon of straight chain or branching, and it can be the multiple substitution values replacements optionally through allowing." alkyl " as used herein example includes, but not limited to methyl, ethyl, propyl group, sec.-propyl, isobutyl-, normal-butyl, the tertiary butyl, isopentyl, and n-pentyl.
As used herein, term " alkylidene group " refers to divalent group, such as " methylene radical ", " ethylidene " and " vinylidene ", and it refers to-the CH of bivalent form respectively 2-,-CH 2-CH 2-, and-CH=CH-.
As used herein, term " aryl " refers to single phenyl ring or fused benzene rings system, and it can be the multiple substitution values replacements optionally through allowing.The example of " aryl " group of effect includes but not limited to phenyl, 2-naphthyl, 1-naphthyl, anthracene and phenanthrene.Preferred aryl rings has 5 to 10 members.
As used herein, the fused benzene rings system that term " aryl " is contained comprises fused polycycle hydrocarbon, that is wherein there is the cyclic hydrocarbon being less than non-accumulated double bond maximum number, such as wherein stable hydrocarbon ring (cycloalkyl is cyclopentyl ring such as) and aromatic ring (aryl is phenyl ring such as) condense to be formed such as group such as indanyl and acenaphthylenyl, and comprise group such as dihydronaphthalene and the naphthane of limiting examples.
Term as used herein " alkoxyl group " refers to group-OR a, wherein R ait is alkyl as defined herein.
Term as used herein " aryloxy " refers to group-OR a, wherein R ait is aryl as defined herein.
" amino " as used herein refers to group-NR ar b, wherein R aand R beach hydrogen naturally.Extraly, " amino replaced " refers to group-NR ar b, wherein R aand R balkyl, arylalkyl or aryl individually separately.As used herein, at R aor R bwhether when hydrogen, above-mentioned group can be called " amino replaced " or, if such as R abe H and R bthat alkyl is then called " alkylamino."
As used herein, term " pharmaceutically acceptable " refers to carrier, thinner, and the salt form of vehicle or the compounds of this invention is compatible with other composition of preparaton and harmless to the recipient of described pharmaceutical composition.
As used herein, term " pharmaceutical composition " refers to the compounds of this invention of optionally pharmaceutically acceptable carrier, thinner or mixed with excipients with one or more.Pharmaceutical composition preferably shows the degree of stability to envrionment conditions, and described degree of stability makes them be suitable for preparation and commercialization intention.
As used herein, term " significant quantity ", " therapeutic dose " and " effective dose " refer to that the amount of the compounds of this invention is enough to cause desired pharmacology or result for the treatment of, thus cause the effective treatment to obstacle.Can show as the treatment of obstacle: to obstacle outbreak or progress and the outbreak of symptom relevant with obstacle or delaying or preventing of progress.Reducing or eliminating of symptom can also be shown as, the reversion of obstacle progress to the treatment of obstacle, and other contribution any to patient's good condition.
Effective dose can depend on following factor and change, such as status of patient, the seriousness of impairment property, and pharmaceutical composition give mode.Usually, in order to give with effective dose, compound can give with the amount being less than 5mg/kg weight in patients.The amount that compound can give is for being less than about 1mg/kg weight in patients to being less than about 100 μ g/kg weight in patients, and about 1 μ g/kg is to being less than 100 μ g/kg weight in patients.The amount that aforesaid effective dose general proxy can give as single dose, or as one or more dosage that can give in 24 hours sections.
The compounds of this invention can comprise the preparation of attested synthetic method by various method.Exemplary general synthetic method is described below, and then specific the compounds of this invention is prepared with working Examples.
In the example be described below, when needing, use blocking group that is responsive or reactive group according to general synthetic chemistry principle.Blocking group handles (T.W.Green and P.G.M.Wuts (1999) Protecting Groups in Organic Synthesis, 3 according to organic synthesis standard method rdedition, John Wiley & Sons, is incorporated to herein by quoting during relevant blocking group).These groups compou nd synthesis facilitate the stage be for a person skilled in the art obvious method removing.To process and the selection of reaction conditions and execution sequence and the preparation of the compounds of this invention consistent.
The present invention also provides the method and their preparation method of synthesizing as the compound of the intermediate prepared in the compounds of this invention.
Compound can according to the method the be described below raw material easily obtained and reagent preparation.In these reactions, those skilled in the art can be used known but the alternative method do not described in detail herein.
Except as otherwise noted, structure described herein is also intended to comprise the compound that difference is only the atom that there is the ground enrichment of one or more isotropic substance.Comprise such compound in the scope of the invention, it has structure of the present invention but hydrogen atom is replaced with deuterium or tritium, or is replaced with by carbon atom 13c-or 14the carbon of C-enrichment.Such as, deuterium is used for checking the pharmacokinetics of biologically active cpds and metabolism widely.Although chemically angle deuterium and hydrogen similar, there is bond energy between deuterium-carbon bond and hydrogen-carbon bond and bond distance's significant difference in them.Thus, in biologically active cpds, replace with deuterium with regard to hydrogen can cause such compound, it usually retains biological chemistry usefulness and selectivity, but the absorption that performance is significantly different compared with it does not contain isotopic counterpart, distribution, metabolism and/or secretion (ADME) characteristic.Thus deuterium replaces pharmaceutical efficacy, safety and/or the tolerance that can cause the improvement of some biologically active cpds.
The compounds of this invention can with more than a kind of form crystallization, and this feature is called polymorphism, and described polymorphic forms (" polymorphic ") belongs within the scope of the invention.Polymorphism usually can occur as to the reaction of temperature, pressure or both changes.Polymorphism can also be caused by the change of crystallisation process.Polymorphic can be characterized by various physical features known in the art such as X-ray diffraction spectrum, solubleness and fusing point.
Some in compound described herein contains one or more chiral centre, or can otherwise exist as multiple steric isomer.The scope of the invention comprises the mixture of the mixture of steric isomer and the enantiomorph of purifying or the ground enrichment of enantiomorph ground/diastereomer.Also the independent isomer of the various compound of the present invention is comprised in the scope of the invention, and the mixture balanced wholly or in part arbitrarily.The present invention also comprises the mixture of the isomer that the independent isomer of above-mentioned various compound and one or more chiral centre overturn.
Compound desirably single enantiomorph time, it can obtain like this: stereospecificity is synthesized, the fractionation of the finished product or arbitrarily intermediate easily, or chiral chromatography methods known in the art.The fractionation of the finished product, intermediate or raw material can be undertaken by any suitable method known in the art.See such as, Stereochemistry of Organic Compounds (Wiley-Interscience, 1994).
The present invention includes salt or the solvate of compound described herein, comprise the solvate that it combines such as salt.The compounds of this invention can exist with solvation such as hydrated form and non-solvation form, and whole described form is contained in the present invention.
Usually, but not utterly, salt of the present invention is pharmacy acceptable salt.The salt that term " pharmacy acceptable salt " is contained refers to the non-toxic salt of the compounds of this invention.
The example of suitable pharmacy acceptable salt comprises inorganic acid addition salt such as muriate, bromide, vitriol, phosphoric acid salt, and nitrate; Organic acid addition salt is acetate such as, mutate, propionic salt, succinate, lactic acid salt, oxyacetate, malate, tartrate, Citrate trianion, maleate, fumarate, mesylate, p-tosylate, and ascorbate salt; There is the salt such as aspartate and glutaminate of acidic amino acid; An alkali metal salt such as sodium salt and sylvite; Alkaline earth salt such as magnesium salts and calcium salt; Ammonium salt; Organic basic salt is front three amine salt such as, triethylamine salt, pyridinium salt, picoline salt, dicyclohexyl amine salt, and N, N'-dibenzyl ethylenediamine salt; With alkaline amino acid salt such as lysine salt and arginic acid salt.Salt can be hydrate or alcohol solvent compound in some cases.
Understanding can provide more than a kind of systematic naming method for many organic compound by the technician of organic chemistry filed.Scope of the present invention should not be considered to unclear because compound exists several possible naming conventions.
II. general synthetic method
The technician in organic synthesis field will understand the means that there is multiple generation the compounds of this invention, and produces the means of the all-purpose radioisotopic the compounds of this invention of mark.
A kind of means producing the compounds of this invention are shown in scheme 1 (see synthetic example).Thus Norcamphor (2-norcamphane ketone) can locate alkylation by the technology known by the technical staff in organic synthesis field contiguous carbonyl official.For described conversion, highly basic (such as sodium hydride, sodium alkoxide, sodium amide) is generally used to process ketone to form enolate intermediate, subsequently with alkyl halide or alkyl sulfonate esters process.Under certain conditions, alkylation can use α, ω-alkylene dihalide (such as 1,3-dibromopropane) to carry out, to form screw connection.Although scheme 1 shows form spirocyclobutane (Compound II per), other ring size (such as, spiro cyclopentane) also can in like fashion by using other α, ω-alkylene dihalide to realize.With Wittig (or of equal value) chemical means, carbonyl official can be converted into the outer methylene radical (compound III) of ring subsequently.Under strong acid exists, process exo-methylene compound with prussic acid (or similar reagents, such as thiocyanate-), can be called that the process that Ritter reacts provides corresponding tertiary carboxamido compounds.With hydride reducer, such as lithium aluminum hydride or two (methoxy ethoxy) sodium alanate reduction carboxamido compounds, provides corresponding secondary amine, compound IV.
Alternatively, the 2-norcamphane ketone of replacement can also be used as the raw material of the conversion described in scheme 2.Thus D-camphor and L-camphor (all commercially available) can be converted into the steric isomer of compound V separately.Other ketone raw material can also be used.Such as, homologue two ring [2 of 2-norcamphane ketone, 2,2] pungent-2-ketone, can pass through two rings [2,2,2] hydrogenation of pungent-5-alkene-2-ketone obtains, the latter can be prepared by the program of similar Kozikowski and Schmiesing, J.Org.Chem.48:1000-1007 (1983) those disclosed again, and relevant reaction is incorporated to herein by quoting.Similarly, hydrogenation can press Black and Vogel, 7-oxabicyclo [2.2.1]-5-in heptan alkene-2-ketone (relevant reaction is incorporated to herein by quoting) prepared by the description of Helv.Chim.Acta 67:1612 (1984) is to provide 7-oxabicyclo [2.2.1] heptane-2-ketone.The each similar potential raw material being shown in those conversion of scheme 1 and 2 naturally of these ketone.
Spirocyclopropane official can enough Simmons-Smith and the realization of chemistry similar means.Thus, under zinc-copper even (zinc-copper couple) exists, 3-methylene radical-2-norcamphane ketone and methylene iodide are reacted and spiral shell [two rings [2.2.1] heptane-2 is provided, 1'-cyclopropane]-3-ketone, then can use the reaction described to be translated into the compounds of this invention.Some to be known in document containing compound of spirocyclopropane and to serve as the starting point of synthesis the compounds of this invention.See such as, Gream andPincombe, Aust.J.Chem.27:543-565 (1974), relevant reaction is incorporated to herein by quoting.
Secondary amine such as compound IV and V can be converted into tertiary amine by acid amides and carbamate intermediate.Thus, process described compound successively with tert-Butyl dicarbonate and lithium aluminum hydride and will produce corresponding N-methyl tertiary amine.
Scheme 1
Scheme 2
Also specific radio isotope may be mixed.Such as, N-tri-deuterium methyl or N-tri-tritium methylamine can be produced with lithium deuteride LiD aluminium or lithium-tritide al reducing agent reducing amide and carbamate.Alternatively, producing carbonyl carbon is 11c, 13c or 14the acid amides of C atom or carbamate, return generation with lithium aluminium hydride reduction subsequently and be mixed with respectively 11c, 13c or 14the amine of C atom.Specifically radioisotopic mixing is usually wish preparing in compound, and described compound is ready to use in diagnostic environment (such as preparation) or function and metabolism research.
III. pharmaceutical composition
Although the compounds of this invention may be given with loose active chemical form, still preferably give compound with pharmaceutical composition or preparaton form.Thus on the one hand the present invention includes pharmaceutical composition, it comprises one or more formula I and/or its pharmacy acceptable salt and one or more pharmaceutically acceptable carrier, thinner or vehicle.Another aspect, the invention provides the method for pharmaceutical compositions, comprises one or more formula I and/or its pharmacy acceptable salt and one or more pharmaceutically acceptable carrier, thinner or mixed with excipients.
The mode giving the compounds of this invention can change.The compounds of this invention preferred oral gives.Preferred pharmaceutical compositions for oral administration comprises tablet, capsule, caplet, syrup, solution and suspension.Pharmaceutical composition of the present invention can provide with modified release dosage form such as timed-release tablets and capsule formulation form.
Pharmaceutical composition can also give via injection, that is through intravenously, through intramuscular, through subcutaneous, in intraperitoneal, intra-arterial, sheath and intracerebral ventricle injection.Intravenous administration is preferred injecting method.Suitable injection carrier is well known to those skilled in the art, comprises the salt solution of 5% glucose solution, salt solution and phosphoric acid buffer.
Each preparaton can also give with other means such as rectal administration.Preparaton such as suppository for rectal administration is well known to those skilled in the art.Described compound can also be given by suction, such as aerosol form; Local gives, such as lotion form; Give through skin, such as use transdermal patch (such as, by the commercially available technology from Novartis and Alza Corporation), powder injection, or cheek, sublingual or Nasal absorption.
Pharmaceutical composition can be formulated as unit dosage form, or multiple doses or subunit's dosage form.
The administration of pharmaceutical composition described herein can be interrupted, or gradual change, continuous print, constant or controlled speed administration.Pharmaceutical composition can give to warm-blooded animal, such as Mammals such as mouse, rat, cat, rabbit, dog, pig, ox or monkey; But advantageously give people.In addition, the opportunity and the number of times that give pharmaceutical composition every day can change.
The compounds of this invention may be used for treating various obstacle and illness, and can thus be used for the treatment of or prevent other suitable therapeutic combinations various of those obstacles or illness to use.Thus one embodiment of the present invention comprise the compounds of this invention that administration and other therapeutic compound combine.Such as, the compounds of this invention can use with following agent combination: other NNR part (such as varenicline), the allosteric modulators of NNRs, antioxidant (such as radical scavenger), antiseptic-germicide (such as penicillin antibiotic), antiviral agent (such as nucleoside analog, as zidovudine and acyclovir), anticoagulation (such as warfarin), anti-inflammatory agent (such as NSAIDs), febrifuge, anodyne, narcotic (such as operation), acetylcholinesterase depressant (such as E2020 and lycoremine), antipsychotic drug (such as haloperidol, leoponex, olanzapine and Quetiapine), immunosuppressor (such as ciclosporin and methotrexate), neuroprotective, steroid (such as steroid hormone class), reflunomide (such as dexamethasone, prednisone and hydrocortisone), VITAMIN, mineral, healthcare products, thymoleptic (such as imipramine, fluoxetine, paroxetine, escitalopram, Sertraline, Venlafaxine and duloxetine), anxiolytic (such as alprazolam and buspirone), anticonvulsive drug (such as Phenytoin Sodium Salt and gabapentin), vasodilator (such as Prazosin and Virga), mood stabilizer (such as valproate and Aripiprazole), anticarcinogen (such as antiproliferative), hypotensive agent (such as atenolol USP 23, clonidine, amlodipine, verapamil and Olmesartan), caccagogue, manure bate, hydragog(ue) (such as Furosemide), spasmolytic (such as Dicycloverine), anti-dyskinesia agent, with anti-ulcerative drug (such as esomeprazole).The combination of above-mentioned forms of pharmacologically active agents can give together or dividually, and when separately giving, administration side by side or in turn can be carried out with any order.Select compound or the amount of reagent and the relative timings of administration so that the result for the treatment of desired by realizing.The combination medicine-feeding of the compounds of this invention and other therapeutical agent can with following form combination medicine-feeding simultaneously: (1) comprises the unit pharmaceutical composition of both compounds; Or (2) respectively comprise the pharmaceutical composition separated of one of described compound.Alternatively, combination can separately give in a sequential manner, wherein first gives a kind of therapeutical agent and then gives the second therapeutical agent.Described order of administration can be separated by of short duration or very long in time.
Another aspect of the present invention comprises combination treatment, and the compounds of this invention comprised to snibject's treatment or prevention significant quantity comprises chemotherapy, radiotherapy, gene therapy or immunotherapy with one or more other therapies.
IV. the method for pharmaceutical composition is made
The compounds of this invention can be used in prevention or treatment has proposed or shown various illness or the obstacle that other type nicotinic compounds can be used as therapeutical agent, such as CNS obstacle, inflammation, the Inflammatory response relevant with bacterium and/or virus infection, pain, metabolism syndrome, autoimmune disorders, habituation, other obstacle that is fat or that describe in further detail herein describes.This compound can also be used as diagnostic reagent (in vitro and in vivo).Described treatment and other instruction are described in the reference such as listed in advance herein, comprise the people such as Williams, Drug News Perspec.7 (4): 205 (1994), the people such as Arneric, CNS Drug Rev.1 (1): 1-26 (1995), the people such as Arneric, Exp.Opin.Invest.Drugs 5 (1): 79-100 (1996), the people such as Bencherif, J.Pharmacol.Exp.Ther.279:1413 (1996), the people such as Lippiello, J.Pharmacol.Exp.Ther.279:1422 (1996), the people such as Damaj, J.Pharmacol.Exp.Ther.291:390 (1999), the people such as Chiari, Anesthesiology 91:1447 (1999), Lavand ' homme andEisenbach, Anesthesiology 91:1455 (1999), the people such as Holladay, J.Med.Chem.40 (28): 4169-94 (1997), the people such as Bannon, Science 279:77 (1998), PCT WO94/08992, PCT WO 96/31475, PCT WO 96/40682, with the U.S. Patent number 5 of the people such as Bencherif, 583, 140, the U.S. Patent number 5 of the people such as Dull, 597, 919, the U.S. Patent number 5 of the people such as Smith, 604, the U.S. Patent number 5 of the people such as 231 and Cosford, 852, 041.
CNS obstacle
Compound and pharmaceutical composition thereof are used for the treatment of or prevent various CNS obstacle, comprise neurodegenerative disease, neuropsychiatric disorders, neurological and habituation.Compound and pharmaceutical composition thereof can be used for treatment or prevention age related and incoherent cognition is low and dysfunction; Attention disorders and dementia, comprise those that cause due to infective agent or metabolism disorder; Neuroprotective is provided; Treatment is fainted from fear and multiple infarction of brain; Treatment affective disorder, forces and Addictive Behaviors; Pain relieving is provided; Control inflammation, such as mediated by cytokine and nf kappa B those; Treatment inflammatory conditions; Pain relief is provided; Infect with treatment, be used for the treatment of bacterium, fungi and virus infection as infection agent.The compounds of this invention and pharmaceutical composition can be used for treat or prevention obstacle, especially comprise in disease and illness: the age related memory impairment (AAMI), mild-cognitive impairement (MCI), age related cognitive declines (ARCD), dementia senilis, Early onset Alzheimer, senile dementia, Alzheimer type is dull-witted, Alzheimer, cognitive disease damage is without dull-witted (CIND), Lewy body is dull-witted, HIV-is dull-witted, AIDS chronic brain syndrome, vascular dementia, Down syndrome, head trauma, traumatic brain injury (TBI), dementia pugilistica, Creutzfeldt-Jakob disease and Puli's protein disease, palsy, central authorities' ischemic, periphery ischemic, distractibility obstacle, Attention deficit hyperactivity disorder, dislexia, schizophrenia, schizophreniform disorder, schizoaffective disorders, cognition dysfunction in schizophrenia, cognition in schizophrenia is low, parkinsonism comprises Parkinson's disease, parkinsonism after encephalitis, parkinsonism-Gaum is dull-witted, frontotemporal dementia Parkinson type (FTDP), pager's disease, Niemann-pager's disease, Huntington Chorea, Huntington Chorea, dyskinesia, tardive dyskinesia, spasm dystonia, hyperkinesis, stein-leventhal syndrome, property paresis on Progressive symmetric erythrokeratodermia core, restless legs syndrome, Creutzfeldt-Jakob disease, multiple sclerosis, amyotrophic lateral sclerosis (ALS), motor neuron (MND), multiple system atrophy (MSA), corticobasal degeneration, guillain-Barre syndrome (GBS), with chronic inflammatory demyelinating polyneuropathy (CIDP), epilepsy, the main nocturnal frontal lobe epilepsy of euchromosome type, mania, anxiety, depressed, comprise major depressive disorder (MDD), premenstruum haves the fidgets, panic disorder, Bulimia nerovsa, anorexia, narcolepsy, excess daytime somnolence, bipolar affective disorder, generalized anxiety disorder, compulsive disorder, indignation outburst, carry out obstacle, oppositional defiant disorder, Tourette syndrome, autism, medicine and alcohol addiction, tobacco addiction, thus be used as smoking stopping, mandatory overfeeding and sexual dysfunction medicine.
Cognitive disease damage or dysfunction can be relevant with psychiatric disorders or illness, such as schizophrenia and other mental disorder, include but not limited to mental disorder, schizophreniform disorder, schizoaffective disorders, delusional disorder, of short duration mental disorder, shared mental disorder, with the mental disorder due to general medicine illness, dementia and other cognitive disorder, include but not limited to mild-cognitive impairement, dementia senilis, Alzheimer, senile dementia, Alzheimer type is dull-witted, Age-Associated Memory defect, Lewy body is dull-witted, vascular dementia, AIDS chronic brain syndrome, dislexia, parkinsonism comprises Parkinson's disease, cognitive disease damage and Parkinson's disease dementia, the cognitive disease damage of multiple sclerosis, the cognitive disease damage caused by traumatic brain injury, due to the dementia of other general medicine illness, anxiety disorder, include but not limited to Panic disorder without agoraphobia, Panic disorder with agoraphobia, without the agoraphobia of panic disorder medical history, specific phobia disease, social phobia, compulsive disorder, posttraumatic stress disorder, acute stress disorder, generalized anxiety disorder and the generalized anxiety disorder due to general medicine illness, affective disorder, include but not limited to major depressive disorder, evil mood mental disorder, two-way type is depressed, two-way type mania, two-way type I obstacle, with manic relevant depression, depressed or mixing shows effect, two-way type II obstacle, cycloophrenia obstacle, with the affective disorder due to general medicine illness, somnopathy, include but not limited to somnopathy, severe insomnia disease, severe hypersomnia, narcolepsy, parasomnia obstacle, nightmare obstacle, night terror and somnambulism obstacle, mental retardation, learning disorder, motor skill disorder, communication disorders, PDD, distractibility and disruptive behavior disorder, distractibility obstacle, Attention deficit hyperactivity disorder, infancy, Childhood or the feeding of adult and eating disorder, tic disorder, remove obstacles, the obstacle that material is relevant, include but not limited to substance depilatory, substance abuse, Substance Intoxication, material is given up, the obstacle that alcohol is relevant, the obstacle that amphetamine or amphetamine analogue are correlated with, the obstacle that caffeine is relevant, the obstacle that hemp is relevant, the obstacle that Cocaine is relevant, the obstacle that fantasy is relevant, the obstacle that inhalation is relevant, the obstacle that Nicotine is relevant, the obstacle that opioid is relevant, the obstacle that phencyclidine or the similar thing of phencyclidine are correlated with, and tranquilizer, the obstacle that soporific or anxiolytic are correlated with, personality disorder, include but not limited to Obsessive-compulsive disorder and impulse control disorder.
Cognitive performance can be evaluated by the cognitive grade of confirmation, the cognitive subscale (ADAS-cog) of such as Alzheimer opinion rating.The intensity of variation according to above-mentioned grade measuring patient can be comprised in a kind of measurement improving the validity in cognition to the compounds of this invention.
Take into account and force and Addictive Behaviors, the compounds of this invention can use the therapy as nicotine addiction, comprise and stop agent as smoking, with for other reward of brain obstacle, such as substance abuse comprises alcohol addiction, illegal medicine and prescription drugs habituation, eating disorder, comprises fat and behavior habituation, such as other similar behavior expression of gambling or habituation.
Above-mentioned condition and obstacle are such as at American Psychiatric Association:Diagnostic and Statistical Manual of Mental Disorders, Fourth Edition, Text Revision, Washington, DC, American Psychiatric Association, 2000 have and discuss in detail further.Can also with reference in this handbook to the more details of the symptom relevant with substance migration, abuse and dependence and diagnostic characteristic.
Inflammation
It is known that the neural system mainly scale of vagus nerve by suppressing the release of scavenger cell tumour necrosis factor (TNF) to regulate innate immune response.This physiological mechanism be called " cholinergic drug anti-inflammatory pathways " (see, such as, Tracey, " The Inflammatory Reflex, " Nature420:853-9 (2002)).Excessive inflammation and tumour necrosis factor synthesis cause morbidity and even death in various disease.These diseases include but not limited to endotoxemia, rheumatoid arthritis, osteoarthritis, psoriasis, asthma, atherosclerosis, idiopathic pulmonary fibrosis, and inflammatory bowel.
Chronic and acute inflammation can be included but not limited to by the inflammatory conditions giving compounds for treating described herein or prevention, psoriasis, endotoxemia, gout, acute pseudogout, acute gouty arthritis, sacroiliitis, rheumatoid arthritis, osteoarthritis, allograft rejection, chronic transplanting rejection, asthma, atherosclerosis, monokaryon-phagocytic cell dependency lung injury, idiopathic pulmonary fibrosis, atopic dermatitis, chronic obstructive pulmonary disease, adult respiratory distress syndrome, acute chest syndrome in sickle cell disease, inflammatory bowel, irritable bowel syndrome, comprise the dominant irritable bowel syndrome of diarrhoea, Crohn's disease, ulcer, ulcerative colitis, acute cholangitis, aphthous stomatitis, emaciation, cryptitis, glomerulonephritis, systemic lupus erythematosus, thrombosis, and graft-vs-host reaction.
The Inflammatory response relevant with bacterium and/or virus infection
It is relevant with the natural response of health to bacterium or virus and/or toxin that many bacteriums and/or virus infection and toxin form the side effect brought.As discussed above, health usually involves the TNF and/or other cytokine that produce significant quantity to the response infected.The overexpression of these cytokines can cause remarkable injury, such as septic shock (when bacterium is septic), endotoxin shock, urosepsis, virus pneumonia and toxic shock syndrome, TSS.
The expression of cytokine is mediated by NNRs, and can by giving these receptor stimulants or partial agonist suppresses.Therefore, can be used for making to minimize with bacteriological infection and viral relevant with fungi infestation Inflammatory response as the agonist of these acceptors or those compounds described herein of partial agonist.The example of described bacteriological infection comprises anthrax, sausage poisoning and Sepsis.Some in these compounds can also have antimicrobial property.In addition, described compound can be used in treating raynaud's disease, that is the Painful peripheral vasoconstriction of virus induction.
These compounds can also be used as adjuvant therapy and existing therapy combines to administer bacterium, virus and fungi infestation; These compounds are such as microbiotic, antiviral drug and antifungal drug.Toxinicide can also be used for being bonded to the toxin of infectious substance generation and the toxin of permission combination does not produce Inflammatory response by health.Antitoxic example is disclosed in the U.S. Patent number 6,310,043 of the people such as such as Bundle.Can be effective to bacterium and other toxin other reagent effective, and their result for the treatment of can by supplementing with compound Combined Preparation described herein.
Pain
Compound can be given to treat and/or prevent pain, comprise acute pain, Neuropathic pain, inflammatory pain, neuropathic pain and chronic pain.Compound can be combined the possibility (such as morphine restraining therapy) minimizing opiate addiction with opiates.The analgesic activities of compound described herein can be illustrated in the model of lasting inflammatory pain and the neuropathic pain of being undertaken by the description of US publication application number 20010056084A1 (people such as Allgeier) (the mechanical hyperalgesia enhanced sensitivity in the mechanical hyperalgesia enhanced sensitivity such as, in the complete Freund adjuvant rat model of inflammatory pain and the mouse Partial sciatic nerve ligation model in neuropathic pain).
Analgesic effect is suitable for treating various origin or etiologic pain, especially inflammatory pain and relevant hyperalgesia is treated, neuropathic pain and relevant hyperalgesia, chronic pain (such as, severe chronic pain, postoperative pain and comprise Cancer-Related pain with various illness, stenocardia, kidney or biliary colic, menstruation, migraine, and gout).Inflammatory pain can originate from different, comprises sacroiliitis and rheumatoid disease, tendon synovitis and vasculitis.Neuropathic pain comprises trigeminal nerve or herpes neuralgia, and neuropathy is diabetic neuropathy pain such as, cusalgia, pain in the back and deafferentation syndromes such as brachial plexus avulsion.
Neovascularization
Such as can treat or prevent feature to be the illness of undesirable neovascularization or vasculogenesis by the suppression of the neovascularization giving the antagonist (or partial agonist of some dosage) of nAChR.It is that inflammatory vascular generates and/or those of ischemic inducibility vasculogenesis that described illness can comprise feature.The neovascularization relevant with tumor growth can also be suppressed by those compounds described herein of the antagonist or partial agonist that serve as nAChR.
The specific antagonistic action of nAChR reduces replys inflammation, ischemic and neoplastic vasculogenesis.The guidance taking into account the suitable animal model system evaluating compound described herein can see such as Heeschen, C. people is waited, " A novel angiogenic pathway mediated bynon-neuronal nicotinic acetylcholine receptors, " J.Clin.Invest.110 (4): 527-36 (2002).
SCLC can be comprised with the representative tumor type of compounds for treating described herein, NSCLC, ovarian cancer, pancreas cancer, mammary cancer, colorectal carcinoma, the rectum cancer, lung cancer, oropharynx cancer, hypopharyngeal cancer, the esophageal carcinoma, cancer of the stomach, pancreas cancer, liver cancer, carcinoma of gallbladder, cholangiocarcinoma, carcinoma of small intestine, urethral carcinoma, kidney, bladder cancer, urothelial cancer, Female Reproductive Tract Cancer, cervical cancer, uterus carcinoma, ovarian cancer, choriocarcinoma, gestational trophoblastic disease, male genital cancer, prostate cancer, carcinoma of seminal vesicle, carcinoma of testis, blastoma, internal secretion gland cancer, thyroid carcinoma, adrenal carcinoma, pituitary carcinoma, skin carcinoma, vascular tumor, melanoma, sarcoma, Bone and soft tissue sarcoma, Kaposi sarcoma, cerebral tumor, neural tumor, eye neoplasms, meningeal tumor, astrocytoma, glioma, glioblastoma multiforme, retinoblastoma, neuroma, neuroblastoma, schwannoma, meningioma, pernicious solid tumor (the such as leukemia caused of hematopoiesis, chlorosarcoma, plasmoma and mycosis fungoides and cutaneous T-cell lymphomas/leukemic patch and tumour), with the solid tumor that lymphoma causes.
Compound can also be combined with the anticancer therapy thing of other form and give, and comprises and antineoplastic agent known in the art such as cis-platinum, Zorubicin, daunomycin etc. and/or the administration of anti-VEGF (vascular endothelial growth factor) agents.
Compound can give thus its target tumor place in the above described manner.Such as, compound can give being conjugated in the microballoon of various antibody of the tumour that to be led by particulate, particulate or liposome.Extraly, compound can be present in microballoon, particulate or liposome, and its size is suitable for by artery and vein but rests in the capillary bed of tumour and to give tumour partly by compound.Described drug delivery device is known in the art.
Other obstacle
Except treatment CNS obstacle, inflammation and neovascularization and pain, the compounds of this invention can also be used for preventing or treat NNRs in some other illness, disease and obstacle of wherein working.Example comprises autoimmune disorders such as lupus, the obstacle relevant with release of cytokines, infect the emaciation of secondary (such as, occur in AIDS, in the syndrome that AIDS is relevant and tumorigenesis), fat, pemphitis, the urinary incontinence, bladder hyperactivity (OAB), diarrhoea, constipation, retinal diseases, transmissible disease, myasthenia, Eaton-Lambert syndrome, hypertension, preeclampsia, osteoporosis, vasoconstriction, vasodilation, arrhythmia, type i diabetes, type ii diabetes, Bulimia nerovsa, anorexia and sexual dysfunction, and those indications disclosed in PCT application WO 98/25619.The compounds of this invention can also be given to faint from fear those of such as epilepsy syndromes and treatment illness such as syphilis and Creutzfeldt-Jakob disease with treatment.
The compounds of this invention can be used for treating bacteriological infection and dermatological conditions, such as pemphigus foliaceus, pemphigus vulgaris, and other obstacle, such as acantholysis, wherein there is the autoimmune response with high neuroganglion NNR IgG titers.In these obstacles, and in other autoimmune disorder such as myasthenia gravis, the fab fragment of antibody is bonded to NNR acceptor (crosslinked 2 acceptors), this induction internalization and degraded.
Diagnostic uses
Compound can be used in diagnosis composition such as probe, when particularly being modified to comprise suitable marker.For this intention, the compounds of this invention most preferably uses radioisotopic moieties 11c marks.
The compound given can detect with position emission scan art (PET).Wish high specific activity with unsaturation concentration by select receptor subtype visual.The dosage that gives generally provides high-contrast image under toxicity range.Compound is expected can with non-toxic level administration.The determination of dosage is carried out in isotopic labeling imaging field mode known to the skilled.See such as, the U.S. Patent number 5,969,144 of the people such as London.
Compound can give with known technology.See such as, the U.S. Patent number 5,969,144 of the people such as above-mentioned London.Compound can give in the preparaton composition mixing other composition, and other composition described is such as preparing those types of the composition of diagnosis composition.Carry out the useful compound of the present invention most preferably to use with high purity.See the U.S. Patent number 5,853,696 of the people such as Elmalch.
After compound being given experimenter (such as human experimenter), the existence of compound in experimenter can by proper technology imaging and quantitative with point out its exist, amount and function.Except the mankind, compound can also give animal such as mouse, rat, dog and monkey.SPECT and PET imaging can be carried out with technology and equipment suitable arbitrarily.See people such as Villemagne, the people such as In:Arneric (Eds.) Neuronal Nicotinic Receptors:Pharmacology andTherapeutic Opportunities, the U.S. Patent number 5 of the people such as 235-250 (1998) and Elmalch, 853,696, be incorporated to herein each via quoting, for open representational imaging technique.
V. synthetic example
Embodiment 1: external form-N, 3-dimethyl spiral shell [two rings [2.2.1] heptane-2,1'-tetramethylene]-3-amine
To 2-norcamphane ketone (Norcamphor) (16.0g, 145mmol) and 1,3-dibromopropane (203mmol, 20.7mL; Diethyl ether (450mL) solution 41.1g) adds sodium amide (363mmol, 14.8g), and mixture is stirred 24 hours under reflux.Mixture is inclined in 200mL frozen water, be separated organic layer.Water layer 200mL ether extracts.The concentrated ether extract through merging, holds in the palm vacuum distillating liquid resistates at 60-100 DEG C in 10-20, obtains the not straight product of 14g.Be dissolved in 150mL hexane, with water (150mL) solution stirring 5 hours of potassium permanganate (12.0g, 75.9mmol).Two-phase mixture is filtered through bed of diatomaceous earth, then uses hexane (100mL) to wash.Be separated hexane layer, water layer 600mL hexane extraction.Merge hexane layer, concentrated, purifying on a silica gel column, with 10-40% ether/hexane wash-out, obtain spiral shell [two rings [2.2.1] heptane-2,1'-tetramethylene]-3-ketone (scheme 1 Compound II per) (6.1g, 28% yield), be oily matter. 1H NMR(CDCl 3，400MHz)：δ2.55-2.49(m，2H)，2.18-2.08(m，2H)，2.00-1.58(m，7H)，1.49-1.36(m，3H)；LCMS(m/z)：151(M+1)。
At-78 DEG C, to (methyl) triphenyl phosphonium bromide (49.9mmol, anhydrous tetrahydro furan (THF) (100mL) solution 18.2g) adds n-Butyl Lithium (the 2.5M hexane solution of 46.5mmol, 18.6mL).Stir the mixture 30 minutes at-78 DEG C.Spiral shell [two rings [2.2.1] heptane-2,1'-tetramethylene]-3-ketone (5.00g, 33.3mmol) is added to this mixture.Gained mixture is stirred 20 hours in envrionment temperature.Add hexane (300mL), filtering mixt.Concentrated filtrate, resistates is at 80g silica column purification, and with Hex, obtaining 3-methylene radical spiral shell [two rings [2.2.1] heptane-2,1'-tetramethylene] (scheme 1 compound III) (3.7g, 75%), is oily matter. 1H NMR(CDCl 3，400MHz)：δ4.82(s，2H)，2.63(brs，1H)，2.22(brs，1H)，2.05-1.78(m，6H)，1.63-1.52(m，1H)，1.48-1.34(m，3H)，1.21-1.12(m，2H)。
At 50 DEG C in 15 minutes, the suspension to 3-methylene radical spiral shell [two rings [2.2.1] heptane-2,1'-tetramethylene] (2.10g, 14.2mmol) and potassium sulfocyanate (14.2mmol, 1.39g) slowly adds sulfuric acid (1.40g; Water (0.52mL) solution 14.3mmol).Solution stirs 5.5 hours at 85 DEG C.Solution is cooled to envrionment temperature, with toluene (20mL) dilution, washs according to priority with water (20mL) and saturated aqueous solution of sodium bicarbonate (10mL).Collect toluene layer, dry on anhydrous sodium sulfate, filter.Add two (methoxy ethoxy) sodium alanate (the 65-70% toluene solution of 28mmol, 7.9mL) to filtrate, gained mixture stirs 2 hours at 85 DEG C.Mixture is cooled to 0 DEG C, slowly adds in batches and drip 3N aqueous NaOH (3mL) and 5% clorox (15mL) mixture.Separation of methylbenzene layer, washs with water (30mL).Then 1N aqueous hydrochloric acid (2x 10mL) extracting toluene layer is used.Discard toluene layer, add 10% aqueous NaOH and make the hcl as extraction agent thing through merging be alkalescence (pH10).Alkaline aqueous mixture ether (2x 30mL) extracts.Collect ether extract, concentrated, by silica gel column chromatography purifying, with 0-40%CMA (chloroform: methyl alcohol: 30% ammoniacal liquor, 9:1:0.1)/chloroform wash-out, obtain external form-N, 3-dimethyl spiral shell [two rings [2.2.1] heptane-2,1'-tetramethylene]-3-amine (0.38g, 15% yield) (scheme 1 compound IV), be oily matter.This oily matter is dissolved in 5mL methylene dichloride, cools in ice bath, merge with the 6M aqueous hydrochloric acid of 2mL.Enriched mixture, vacuum-drying, obtains hydrochloride. 1H NMR(D 2O，400MHz)：δ2.41(s，3H)，2.24-2.18(m，2H)，1.98-1.90(m，1H)，1.82-1.74(m，1H)，1.67-1.58(m，2H)，1.52-1.11(m，8H)，0.95(s，3H)；LCMS(m/z)：180(M+1)。
External form stereochemistry is determined by NMR.
Embodiment 2: the chiral chromatography of external form-N, 3-dimethyl spiral shell [two rings [2.2.1] heptane-2,1'-tetramethylene]-3-amine is separated
By external form-N, 3-dimethyl spiral shell [two rings [2.2.1] heptane-2,1'-tetramethylene]-3-amine (2.0g) is dissolved in 20mL acetonitrile, be injected on chiral column with 0.2mL and be separated (chirality Pak AD-H, 5 microns, 250x 20cm), acetonitrile/Virahol (95:5) eluant solution of the diethylamine with 0.2%, flow velocity 10mL/min.Concentrated peak 1 (early wash-out) and peak 2 (late wash-out) containing fraction dividually.Two kinds of resistatess are dissolved in 10mL methylene dichloride individually, with the process of 2mL 6N aqueous hydrochloric acid, are concentrated into dry.These hydrochloride products are heavy 0.74g (peak 1) and 0.48g (peak 2) respectively.
Embodiment 3:N, 3-dimethyl spiral shell [two rings [2.2.1] heptane-2,1'-pentamethylene]-3-amine
Diethyl ether (700mL) solution to 2-norcamphane ketone (25.0g, 227mmol) and Isosorbide-5-Nitrae-dibromobutane (68.0g, 317mmol) adds sodium amide (23.1g, 567mmol).This mixture heats 24 hours under reflux, and cooling, inclines in 200mL ice-water.Collected organic layer, water layer 200mL diethyl ether.The concentrated diethyl ether thing through merging, resistates in 7-15 holder distillation, obtains the not straight product of 19g at 65-80 DEG C.Be dissolved in hexane (500mL), stirred 5 hours with moisture potassium permanganate (30g, 0.19mol, 500mL).Filtering mixt, collects hexane layer.Water layer 600mL hexane extraction.The concentrated hexane layer through merging, resistates is purifying on a silica gel column, and with 5-15% ethyl acetate/hexane wash-out, obtaining spiral shell [two rings [2.2.1] heptane-2,1'-pentamethylene]-3-ketone (12.6g, 33.8%), is oily matter. 1H NMR(CDCl 3，400MHz)：δ2.56(d，J＝5.1Hz，1H)，2.24(bs，1H)，1.44-1.88(m，14H)；LCMS(m/z)：165(M+1)。
At-78 DEG C, THF (100mL) solution to (methyl) triphenyl phosphonium bromide (17.6g, 48.4mmol) adds n-Butyl Lithium (18.1mL 2.5M THF solution, 45mmol), stirs the mixture 30 minutes.Spiral shell [two rings [2.2.1] heptane-2,1'-pentamethylene]-3-ketone (5.30g, 32.3mmol) is added, envrionment temperature stirring reaction 18 hours to this mixture.Add hexane (200mL), filtering mixt.Concentrated filtrate, resistates is at 80g purified on silica column, and with Hex, obtaining 3-methylene radical spiral shell [two rings [2.2.1] heptane-2,1'-pentamethylene] (4.80g, 91.7%), is oily matter.
At 50 DEG C, by sulfuric acid (1.61mL, 2.96g, 30.2mmol) slowly add to 3-methylene radical spiral shell [two rings [2.2.1] heptane-2,1'-pentamethylene] suspension of (4.80g, 29.6mmol) and potassium sulfocyanate (2.96g, 30.2mmol).Then, reaction mixture is stirred 5.5 hours at 85 DEG C, is cooled to envrionment temperature, with toluene (30mL) dilution, wash with water (20mL), use saturated aqueous solution of sodium bicarbonate (10mL) to wash subsequently.Toluene layer is dry on anhydrous sodium sulfate, filters.Two (methoxy ethoxy) sodium alanate (40% toluene solution, 2 equivalents) is added, 85 DEG C of stirring reactions 2 hours to filtrate.Reaction is cooled to 0 DEG C, slowly adds 5% moisture clorox (35mL) solution of (interval dropping) 3N aqueous NaOH (20mL).Separation of methylbenzene layer, washs with water (30mL).Then, by toluene layer 1N aqueous hydrochloric acid (2x 10mL) extraction, toluene layer is discarded.Adding 10% aqueous NaOH makes aqueous hydrochloric acid layer in alkalescence (pH 10), uses diethyl ether.Concentrated diethyl ether thing, resistates is by silica gel column chromatography purifying, with 0-40%CMA (chloroform: methyl alcohol: 30% moisture ammoniacal liquor, 9:1:0.1)/chloroform, obtain N, 3-dimethyl spiral shell [two rings [2.2.1] heptane-2,1'-pentamethylene]-3-amine (1.2g, 53%), be oily matter. 1H NMR(CDCl 3，400MHz)：δ2.28(s，3H)，2.24(bs，1H)，1.84-1.76(m，2H)，1.73-1.68(m，1H)，1.62-1.52(m，4H)，1.48-1.24(m，7H)，1.09-1.05(m，1H)，1.04(s，3H)；LCMS(m/z)：194(M+1)。
Embodiment 4: the general procedure of spiral shell [two rings [2.2.1] heptane-2,1'-the tetramethylene]-3-amine that preparation N-replaces
Spiral shell [two rings [2.2.1] heptane-2,1'-the tetramethylene]-3-amine that some N-replaces can be prepared like this: by spiral shell [two rings [2.2.1] heptane-2,1'-tetramethylene]-3-ketone body powder amination.Following program is exemplary, and it uses methylamine and provides N-methylspiro [two rings [2.2.1] heptane-2,1'-tetramethylene]-3-amine trifluoroacetate.Utilization dimethylamine, azetidine carry out reduction amination with tetramethyleneimine with similar.
To spiral shell [two rings [2.2.1] heptane-2,1'-tetramethylene]-3-ketone (0.15g, 1.0mmol) with methylamine (4.0mL 2.0M THF solution, 8.0mmol) 1,2-ethylene dichloride (10mL) solution adds acetic acid (0.2mL) and sodium triacetoxy borohydride (0.85g, 4.0mmol).Envrionment temperature stirring reaction 48 hours, with methylene dichloride (10mL) dilution, with saturated sodium bicarbonate aqueous solution (10mL) washing, concentrated.Resistates is purifying on preparation HPLC, with the mixture wash-out of 0.05% formic acid/water and 0.05% formic acid/acetonitrile.Concentrated selected fraction, is dissolved in methyl alcohol (2mL) by resistates.Add trifluoroacetic acid (0.1mL), enriched mixture, vacuum-drying, obtaining N-methylspiro [two rings [2.2.1] heptane-2,1'-tetramethylene]-3-amine trifluoroacetate (0.088g), is jelly. 1H NMR(CD 3OD，400MHz)：δ3.06-3.02(m，1H)，2.568(s，3H)，2.54(brs，1H)，2.34(brs，1H)，2.02-1.83(m，6H)，1.56-1.42(m，5H)，1.26-1.32(m，1H)；LCMS(m/z)：166(M+1)。
Embodiment 5:(1S, 3R, 4R)-N, 4,7,7-tetramethyl-spiral shell [two rings [2.2.1] heptane-2,1'-tetramethylene]-3-amine hydrochlorate and (1S, 3S, 4R)-N, 4,7,7-tetramethyl-spiral shell [two rings [2.2.1] heptane-2,1'-tetramethylene]-3-amine hydrochlorate
Repeat as raw material the following chemical process making raw material with D-camphor with L-camphor, produce as those the product of enantiomorph described herein.
At 100 DEG C, by D-(+)-camphor (4.40g, 28.9mmol) and sodium amide (2.50g, 61.5mmol), the mixture in toluene (100mL) stirs 30 minutes.Toluene (20mL) solution of 1,3-dibromopropane (31.8mmol, 3.24mL, 6.42g) is added, under reflux reacting by heating 3 hours.Reaction is cooled to envrionment temperature, with water (100mL) washing, dry on anhydrous sodium sulfate, concentrated.Resistates is dissolved in 5% methyl alcohol/methyl chloride (80mL), is cooled to-78 DEG C.Solution leads to ozone until keep blue (~ 10 minutes).Then add dimethyl thioether (2mL), slow warm reaction is to envrionment temperature.Concentrated reaction mixture, resistates is at the upper purifying of silicagel column (40g), with 0-20% ether/Hex, obtain (1S, 4R)-4,7,7-trimethylammonium spiral shell [two rings [2.2.1] heptane-2,1'-tetramethylene]-3-ketone (1.66g, 29.9% yield) is oily matter. 1H NMR(CDCl 3，400MHz)：δ2.26(m，1H)，2.10-1.97(m，5H)，1.85-1.1.66(m，2H)，1.62-1.53(m，1H)，1.47-1.40(m，1H)，1.28-1.19(m，1H)，0.94(s，3H)，0.88(s，3H)，0.75(s，3H)；LCMS(m/z)：193(M+1)。
At 175 DEG C, by (1S, 4R)-4,7,7-trimethylammonium spiral shell [two rings [2.2.1] heptane-2,1'-tetramethylene]-3-ketone (1.60g, 8.32mmol) and the mixture of methane amide (10mL) in formic acid (7mL) stir 72 hours.Reaction mixture is cooled to envrionment temperature, inclines in 200mL ice-water, extract with ether (2x 50mL).Ether extract use water (40mL) washing through merging, dry on anhydrous sodium sulfate, concentrated, obtain N-(4,7,7-trimethylammonium spiral shell [two rings [2.2.1] heptane-2,1'-tetramethylene]-3-base) methane amide (1.55g, 84.2% yield), be jelly.
At 0 DEG C, to N-(4,7,7-trimethylammonium spiral shell [two rings [2.2.1] heptane-2,1'-tetramethylene]-3-base) methane amide (1.50g, THF (40mL) solution 6.78mmol) slowly adds lithium aluminum hydride (27.1mmol, 27.1mL 1.0M THF solution).After addition was complete, by reaction backflow 48 hours.Reaction mixture is cooled to 0 DEG C, adds solid sodium sulfate decahydrate (10g) quencher in batches.After stirring 1 hour, filter this mixture, concentrated filtrate.Resistates at 40g purified on silica column, with 0-100%CMA (chloroform: methyl alcohol: 30% moisture ammoniacal liquor; 9:1:0.1)/chloroform give elutriant, obtains external form-amine product, (1S, 3R, 4R)-N, 4,7,7-tetramethyl-spiral shell [two rings [2.2.1] heptane-2,1'-tetramethylene]-3-amine (0.49g; 35% yield), and inner mold-amine product, (1S, 3S, 4R)-N, 4,7,7-tetramethyl-spiral shell [two rings [2.2.1] heptane-2,1'-tetramethylene]-3-amine (0.30g; 21% yield), be all oily matter.It is dissolved in separately the hydrochloride that two kinds of products are converted into them by 1mL concentrated hydrochloric acid, concentrated, vacuum dried sample.External form-(1S, 3R, 4R)-N, 4,7,7-tetramethyl-spiral shell [two rings [2.2.1] heptane-2,1'-tetramethylene]-3-amine hydrochlorate 1h NMR (D 2o, 400MHz): δ 2.80 (s, 3H), 2.72 (brs, 1H), 2.22-1.91 (m, 4H), 1.84-1.72 (m, 3H), 1.54-1.45 (m, 2H), 1.38-1.31 (m, 1H), 1.10-1.01 (m, 1H), 0.87 (s, 3H), 0.75 (s, 3H), 0.72 (s, 3H); LCMS (m/z): 208 (M+1).Inner mold-(1S, 3S, 4R)-N, 4,7,7-tetramethyl-spiral shell [two rings [2.2.1] heptane-2,1'-tetramethylene]-3-amine hydrochlorate 1h NMR (D 2o, 400MHz): δ 3.12 (brs, 1H), 2.79 (s, 3H), 2.26-2.16 (m, 1H), 2.01-1.85 (m, 3H), 1.78-1.69 (m, 3H), 1.60-1.51 (m, 1H), 1.36-1.24 (m, 2H), 1.08-1.00 (m, 1H), 0.85 (s, 3H), 0.78 (s, 3H), 0.75 (s, 3H); LCMS (m/z): 208 (M+1).
Embodiment 6: general procedure secondary amine being converted into N-methyl tertiary amine
Some N-methyl tertiary amine can be prepared like this: the corresponding secondary amine of reduction amination.Following program is exemplary, and it uses formaldehyde and provides external form-(1S, 3R, 4R)-N, N, 4,7,7-pentamethyl-spiral shell [two rings [2.2.1] heptane-2,1'-tetramethylene]-3-amine hydrochlorate.Similar N-methylation reaction is carried out to various secondary amine.
To (1S, 3R, 4R)-N, 4,7,7-tetramethyl-spiral shell [two rings [2.2.1] heptane-2,1'-tetramethylene]-3-amine (0.10g, methyl alcohol (4mL) solution of 0.48mmol) He 30% water-containing acetal (1mL) added sodium triacetoxy borohydride (0.31g, 1.4mmol), envrionment temperature stirring reaction 16 hours.Reaction, with saturated sodium bicarbonate aqueous solution (30mL) cancellation, extracts with methylene dichloride (2x 30mL).Formic acid (0.2mL) is added the organic extract through merging, rotary evaporimeter is concentrated.Resistates passes through preparative LCMS purifying, with 0.05% formic acid/water and 0.05% formic acid/acetonitrile mixture wash-out.Comprise the fraction of selection, add 10% aqueous NaOH and make it be alkalescence (pH 9), extract with methylene dichloride (2x 30mL).Through the organic extract 0.5mL concentrated hydrochloric acid process merged.This mixture concentrated, vacuum-drying, obtain (1S, 3R, 4R)-N, N, 4,7,7-pentamethyl-spiral shell [two rings [2.2.1] heptane-2,1'-tetramethylene]-3-amine hydrochlorate (0.06g) is white solid. 1H NMR(D 2O，400MHz)：δ3.27(s，3H)，3.19(s，3H)，3.10(s，1H)，2.55-2.28(m，4H)，2.18-1.97(m，3H)，1.80-1.60(m，3H)，1.45-1.36(m，1H)，1.28(s，3H)，1.02(s，3H)，1.01(s，3H)；LCMS(m/z)：222(M+1)。
Embodiment 7:N-methylspiro [two rings [2.2.1] heptane-2,1'-cyclopropane]-3-amine trifluoroacetate
By pure 3-methylene radical-2-norcamphane ketone (8.9g, 73mmol), pure subsequently methylene iodide (8.30mL, 103mmol) add the slurry of the zinc-copper even (9.1g, 57mmol) in diethyl ether (75mL).Heat gained mixture 6 hours under reflux.Add second section zinc-copper even (10g), continue extra 16 hours of backflow.Then, with water (200mL) cancellation reaction, dilute with diethyl ether (200mL).Two-phase mixture is filtered through Celite pad.Be separated organic layer, with 10% aqueous hydrochloric acid (2x50mL) washing, dry on anhydrous magnesium sulfate, concentrated.By resistates by silicagel column, use dichloromethane eluent.Concentrated selected fraction, in 3 holder vacuum distilling resistatess on bottle-p-bottle (bulb-to-bulb) distillation plant, collecting spiral shell [two rings [2.2.1] heptane-2,1'-cyclopropane]-3-ketone (1.6g), is oily matter. 1H NMR(CDCl 3，400MHz)：δ2.71-2.69(m，1H)，2.06(brs，1H)，2.00-1.72(m，3H)，1.66-1.57(m，3H)，1.09-1.00(m，2H)，0.91-0.89(m，1H)，0.80-0.75(m，1H)。
To spiral shell [two rings [2.2.1] heptane-2,1'-cyclopropane]-3-ketone (0.13g, 0.96mmol) with methylamine (4.0mL 2.0M THF solution, 8.0mmol) 1,2-ethylene dichloride (10mL) solution adds acetic acid (0.2mL) and sodium triacetoxy borohydride (0.85g, 4.0mmol), envrionment temperature stirring reaction 48 hours.By reaction methylene dichloride (10mL) dilution, with the washing of saturated sodium bicarbonate aqueous solution (10mL), concentrated.Resistates is purifying on preparation HPLC, with 0.05% formic acid/water and 0.05% formic acid/acetonitrile mixture wash-out.Concentrated selected fraction, is dissolved in methyl alcohol (2mL) by resistates.Add trifluoroacetic acid (0.1mL), enriched mixture, vacuum-drying, obtaining N-methylspiro [two rings [2.2.1] heptane-2,1'-tetramethylene]-3-amine trifluoroacetate (0.005g), is jelly. 1H NMR(CD 3OD，400MHz)：δ2.76(brs，1H)，2.59(s，3H)，1.85-1.82(m，1H)，1.69-1.55(m，6H)，1.34-1.28(m，1H)，0.83-0.78(m，1H)，0.67-0.62(m，1H)，0.58-0.50(m，2H)；LCMS(m/z)：152(M+1)。
Embodiment 8:N, 3-dimethyl spiral shell [two rings [2.2.1] heptane-2,1'-cyclopropane]-3-amine hydrochlorate
Under nitrogen, to such as Gream and Pincombe, (3-methylspiro [two rings [2.2.1] heptan [5] alkene-2 prepared by the description of Aust.J.Chem.27:543-565 (1974) (being incorporated to herein by quoting), 1'-cyclopropane]-3-base) methyl alcohol (20mL) solution of methyl alcohol (9.4g, 57mmol) adds 0.8g 10%Pd/C (soaking).Atmosphere is replaced, in envrionment temperature jolting mixture 4 hours with hydrogen (50psi).Then, reaction is filtered through Celite pad, is then used methanol wash.Concentrated filtrate, producing (3-methylspiro [two rings [2.2.1] heptane-2,1'-cyclopropane]-3-base) methyl alcohol of 9.60g, is white solid (99%).
Solution in the water (30mL) that chromium trioxide (8.0g, 76mmol) in stirring cools in ice bath carefully adds 96% sulfuric acid (6.9mL, 120mmol).Continue cool and stir oxidizing agent solution in ice bath, and by (3-methylspiro [two rings [2.2.1] heptane-2,1'-cyclopropane]-3-base) methyl alcohol (9.5g in 20 minutes sections; Acetone (115mL) solution 57mmol) adds.After addition was complete, stirred reaction mixture is warmed to envrionment temperature in 3 hours simultaneously.Then, by reaction use water (45mL) and ethyl acetate (200mL) dilution.Add sodium bisulfite powder lentamente until brown dissipates and water layer becomes blue.Then phase-splitting, aqueous layer with ethyl acetate (2x 100mL) washs.Merge organic layer, dry on anhydrous magnesium sulfate.Cross and filter siccative, concentrated filtrate, produce green oil thing.Oily matter is column chromatography eluting by silica gel (200g), with 0-50% ethyl acetate/hexane gradient elution.Merge selected fraction, concentrated, producing 3-methylspiro [two rings [2.2.1] heptane-2,1'-the cyclopropane]-3-carboxylic acid of 6.0g, is white solid (58%).
3-methylspiro [two rings [2.2.1] heptane-2 in stirring, 1'-cyclopropane]-3-carboxylic acid (2.8g, 15mmol) with triethylamine (2.6mL, solution in the toluene (70mL) 18mmol) cooled in ice bath adds diphenyl phosphonic acid trinitride (3.5mL, 16mmol).Reaction mixture is warmed to 90 DEG C, stirs 2.5 hours.Then, phenylcarbinol (1.7mL, 16mmol) is added reaction, at 16 hours that 90 DEG C stir the mixture extra.Reaction mixture, concentrated.Resistates is column chromatography eluting by silica gel (60g), with 0-15% ethyl acetate/hexane gradient elution.Merge selected fraction, concentrated, produce the mixture of the material of 1.6g, comprising 3-isocyanato-3-methylspiro [two rings [2.2.1] heptane-2,1'-cyclopropane] and corresponding Benzylcarbamate, is white solid.This mixture is dissolved in anhydrous THF (16mL), cools in ice bath.Slowly add lithium aluminum hydride (8.5mL2.0M THF solution, 17mmol).Reaction is warmed to 55 DEG C and continues 3 hours.Then, in ice bath, cooling reaction, dilutes with diethyl ether (20mL).Careful adds water until gas generation is calmed down, and quencher is reacted.Stir gained sticky white slurry 1 hour in envrionment temperature, salt becomes granulating more during this period.Then, by slurries filtration by Celite pad, filter cake diethyl ether (10mL) then ethyl acetate (10mL) washing.Filtrate through merging extracts with 6M hydrochloric acid (3x4mL).Merge aqueous extract, concentrated on rotary evaporimeter, produce the N of 1.6g, 3-dimethyl spiral shell [two rings [2.2.1] heptane-2,1'-cyclopropane]-3-amine hydrochlorate is white solid (53% yield). 1H NMR(400MHz，D 2O)：δ2.52(s，1H)，2.46(s，3H)，1.72(d，J＝11Hz，1H)，1.49-1.41(m，3H)，1.39-1.32(m，2H)，1.28(d，J＝11Hz，1H)，1.02(s，3H)，0.59-0.51(m，3H)，0.45-0.43(m，1H)；LCMS(m/z)：166(M+1)。
Embodiment 9:N, 3-dimethyl spiral shell [two rings [2.2.2] pungent [5] alkene-2,1'-pentamethylene]-3-amine hydrochlorate and N, 3-dimethyl spiral shell [two rings [2.2.2] octane-2,1'-pentamethylene]-3-amine hydrochlorate
With Kozikowski and Schmiesing, the program (being incorporated to herein by quoting) that J.Org.Chem.48:1000-1007 (1983) describes prepares intermediate two ring [2,2,2] pungent-5-alkene-2-ketone, then N is converted into subsequently, 3-dimethyl spiral shell [two rings [2.2.2] pungent [5] alkene-2,1'-pentamethylene]-3-amine and N, 3-dimethyl spiral shell [two rings [2.2.2] octane-2,1'-pentamethylene]-3-amine.
By vinyl cyanide (79.4g, 1.49mol), 1，3-cyclohexadiene (60g, 0.75mol), and the mixture of quinhydrones (1.1g, 10mmol) to be encapsulated in pipe and 120 DEG C of heating 18 hours.Concentrated gained mixture, silica gel passes through chromatography purification, with ethyl acetate (0.5% to 1%)/petroleum ether mixtures wash-out, 5-cyano group two ring [2 is provided, 2,2] the separable mixture of the isomer (roughly inner mold/external form) of oct-2-ene (64g, 64% yield), is white semi-solid. 1H NMR(300MHz，CDCl 3)δ1.32(m，2H)，1.75(m，3H)，2.04(m，1H)，2.43(m，1H)，2.62(m，1H)，2.78(m，1H)，6.23(m，1H)，6.30(m，1H)； 1H NMR(300MHz，CDCl 3)δ1.28(m，2H)，1.50(m，3H)，1.94(m，1H)，2.68(m，2H)，2.87(m，1H)，6.29(m，1H)，6.44(m，1H)；LCMS(m/z)：134(M+1)。
Pyridine (14.2g in backflow, 0.180mol), phosphorus pentachloride (28.0g, 0.135mol) drip 5-cyano group two ring [2 with the mixture of chloroform (100mL), 2,2] chloroform (50mL) solution of oct-2-ene (12g, 90mmol).Heat gained mixture 15 hours under reflux, cooling, is poured onto on ice.Concentration of organic layers, resistates by chromatography at purified over silica gel, with the mixture wash-out of ethyl acetate (0.5% to 1%)/sherwood oil, 5-chloro-5-cyano group two ring [2,2,2] oct-2-ene (14.3g is provided, 95% yield), be white semi-solid. 1H NMR(300MHz，CDCl 3)δ1.3-1.5(m，3H)，2.02-2.18(m，2H)，2.51(m，1H)，2.72(m，1H)，3.12(m，1H)，6.22(m，1H)，6.41(m，1H)；GCMS(m/z)：167。
5-chloro-5-cyano group two ring [2 in stirring, 2,2] oct-2-ene (65g, 0.39mol) methyl-sulphoxide (500mL) solution of (foregoing routine is for several times carried out in representative) adds potassium hydroxide (87.4g, 1.56mol) and water (30mL).At room temperature stir gained mixture 15 hours, with water (1000mL) dilution, extract with ether (4x500mL).Through the ether extract salt water washing merged, dry on anhydrous sodium sulfate, concentrated.Resistates is by chromatography at purified over silica gel, and with the mixture wash-out of diethyl ether (1% to 5%)/sherwood oil, providing two rings [2,2,2] pungent-5-alkene-2-ketone (23.8g, 50% yield), is white solid. 1H NMR(300MHz，CDCl 3)δ1.53～1.84(m，4H)，2.01～2.02(m，2H)，2.96～2.99(m，1H)，3.11～3.13(m，1H)，6.15～6.21(m，1H)，6.43～6.48(m，1H)；GCMS(m/z)：122。
At-78 DEG C, n-Butyl Lithium (56.5mL 1.6M hexane solution, 90.4mmol) is added anhydrous THF (108mL) solution of Diisopropylamine (11.2mL, 8.05g, 79.6mmol).Mixture is warmed to 0 DEG C, stirs 30 minutes.Solution is cooled to-78 DEG C again, adds two rings [2,2, the 2] pungent-5-alkene-2-ketone (5.00g, 36.2mmol) being dissolved in THF (10mL).-78 DEG C of stirring reactions 30 minutes, then add hexamethyl phosphoric triamide (13.9mL, 14.3g, 79.6mmol) and follow by Isosorbide-5-Nitrae-dibromobutane (4.76mL, 8.59g, 39.8mmol).Reaction mixture is warmed to envrionment temperature, stirs 16 hours, with ammonium chloride saturated aqueous solution (50mL) cancellation, with ether (100mL) dilution, wash with water (3X 50mL).Organic layer is dry on anhydrous sodium sulfate, filters, concentrated.By resistates purifying on 120g silica column, with 100% Hex, 4 times of column volumes, gradient elution is to the hexane/ethyl acetate of 9:1 subsequently.Concentrated selected fraction, produces spiral shell [two rings [2.2.2] pungent [5] alkene-2,1'-pentamethylene]-3-ketone (5.1g; ~ 90%GC/MS is pure), be clear oil thing.Material is not added and is dissolved in anhydrous THF (20mL) with being further purified and being cooled to-78 DEG C.Then add Methyl magnesium bromide (28.6mL 3.0M diethyl ether solution, 85.8mmol), sluggish is warmed to envrionment temperature.Envrionment temperature stirring reaction 18 hours, carefully add ammonium chloride saturated aqueous solution quencher.Reaction is transferred to separating funnel, removing water layer.Organic layers with water (10mL) washes twice, dry on anhydrous sodium sulfate, filters, concentrated.Remaining material (colorless oil) is the mixture of 3-methylspiro [two rings [2.2.2] pungent [5] alkene-2,1'-pentamethylene]-3-alcohol (4.9g) and raw material.
Do not add and be further purified ground, acetic acid (20mL) solution of the sample of above-mentioned firm generation and sodium cyanide (1.93g, 37.8mmol) is merged.This mixture is cooled to 0 DEG C, while this temperature stirs, slowly adds sulfuric acid (20mL).Reaction becomes scarlet after adding each reagent completely, and it is stirred 18 hours in envrionment temperature.Then, it being gone out by adding 100mL shrend, adding 3M aqueous NaOH and making it, in alkalescence (pH 9), to extract with methylene dichloride (4x 50mL).Organic layer through merging is dry on anhydrous sodium sulfate, concentrated, obtains greyish white solid.Solid is dissolved in anhydrous THF (200mL), is cooled to 0 DEG C and remains on this temperature, lithium aluminum hydride (the THF solution of 25.2mL2M, 50.4mmol) solution is slowly added.Reaction is heated to backflow 18 hours, in cooling ice bath, carefully adds the sodium sulfate decahydrate quencher of 5g.Stir gained mixture 30 minutes, filter.Concentrated filtrate, resistates, at 120g purified on silica column, with 0-70%CMA/ chloroform wash-out, provides N, 3-dimethyl spiral shell [two rings [2.2.2] pungent [5] alkene-2,1'-pentamethylene]-3-amine (0.20g, 2.7% yield).This material is dispersed in methylene dichloride (5ml), is converted into HCL salt with the process of 0.5mL 4M Yan Suan dioxane solution, concentrated gained mixture.Amorphous solid is as a result dissolved in methyl alcohol (3mL), precipitates with diethyl ether (3mL).Absorb solvent, will precipitate and grind three times with diethyl ether (3mL).Then, by hydrochloride sample vacuum-drying. 1H NMR(300MHz，CD 3OD)δ1.02(s，3H)，1.19-1.42(m，4H)，1.51-1.64(m，7H)，1.81(m，1H)，2.21(m，2H)，2.73(s，3H)，5.57(dd，J 1＝9Hz，J 2＝3Hz，1H)，6.01(d，J＝6Hz，1H)；LCMS(m/z)：206(M+1)。
N, 3-dimethyl spiral shell [two rings [2.2.2] pungent [5] alkene-2,1'-pentamethylene]-3-amine (80mg, 0.39mmol) is dissolved in methyl alcohol (7.8mL), adds 10%Pd/C (soaking) (41mg).Under this mixture is placed in hydrogen balloon, stir 16 hours in envrionment temperature.Then, reaction mixture is filtered through diatomite, concentrated filtrate, residue N, 3-dimethyl spiral shell [two rings [2.2.2] octane-2,1'-pentamethylene]-3-amine (45mg, 56% yield).Be dissolved in methylene dichloride (3mL), be converted into its hydrochloride with the process of 0.3mL 4M Yan Suan dioxane solution, concentrated gained mixture.Amorphous solid is as a result dissolved in methyl alcohol (3mL), precipitates with diethyl ether (1mL).Absorb solvent, precipitation diethyl ether (3mL) is ground three times.Then, by hydrochloride sample vacuum-drying. 1H NMR(300MHz，CD 3OD)δ0.99(s，3H)，1.31(m，2H)，1.45-1.70(m，10H)，1.85(m，1H)，2.08(m，3H)，2.22-2.35(m，1H)，2.65(s，3H)，3.02(m，1H)；LCMS(m/z)：208(M+1)。
VI. biology test
In the interactional sign at nAChR place
Material and method
The clone .SH-EP1-mankind α 4 β 2 (people such as Eaton, 2003), the SH-EP1-mankind α 4 β 4 (people such as Gentry, 2003) and SH-EP1-α 6 β 3 β 4 α 5 (people such as Grinevich, 2005) clone derive from Dr.Ron Lukas (Barrow Neurological Institute).SH-EP1 clone, PC12, Eagle substratum (the Invitrogen that SH-SY5Y and TE671/RD cell is modified at Dulbecco, Carlsbad, California) in remain on the proliferate stage, above-mentioned substratum is containing 10% horse serum (Invitrogen), 5% foetal calf serum (HyClone, Logan UT), 1mM Sodium.alpha.-ketopropionate, 4mM L-glutaminate.In order to maintain stable transfectant, α 4 β 2 and α 4 β 4 cell culture medium 0.25mg/mL zeocin and 0.13mg/mL hygromycin B are supplemented.For α 6 β 3 β 4 α 5 cell 0.25mg/mL zeocin, 0.13mg/mL hygromycin B, 0.4mg/mL Geneticin, and 0.2mg/mL blasticidin keeps selecting.
Receptors bind is tested
Film is prepared from rat tissue.Rat cortex derives from Analytical Biological Services, Incorporated (ABS, Wilmington, Delaware).Anatomic tissue from female Sprague-Dawley rat, on dry ice freezing and transport.Tissue storage at-20 DEG C until film preparation need.At ice-cold preparative buffer reagent (11mM KCl, the 6mM KH of 10 volumes (weight: volume) 2pO 4, 137mM NaCl, 8mM Na 2hPO 4, 20mM HEPES (free acid), 5mM iodo-acid amide, 1.5mM EDTA, 0.1mM PMSF pH 7.4) in, collected by Polytron (Kinematica GmbH, Switzerland) and the cortex of homogenize 10 rats.At 4 DEG C, by gained homogenize thing in 40,000g centrifugal 20 minutes, by gained granule settling flux in the ice cold water of 20 volumes.After 4 DEG C of incubations 60 minutes, within centrifugal 20 minutes, collect new granule at 4 DEG C in 40,000g.By final granule settling flux in preparative buffer reagent, and be stored in-20 DEG C.In test day, tissue is thawed, in 40,000g centrifugal 20 minutes, and then be suspended in the salt solution of Dulbecco phosphoric acid buffer, pH 7.4 (PBS, Invitrogen), until the ultimate density of 2-3mg protein/mL.With Pierce BCA protein test kit (PierceBiotechnology, Rockford, IL), adopt bovine serum albumin as standard, measure protein concn.
Film is prepared from cloned cell line.Harvested cell in the PBS of ice-cold pH 7.4, then uses Polytron (Kinematica GmbH, Switzerland) homogenize.Homogenize thing is in centrifugal 20 minutes of 40,000g (4 DEG C).By granule settling flux in PBS, measure protein concn with Pierce BCA protein test kit (penetrating biotechnology, Rockford, IL).
To the competition binding of the acceptor in film preparation.With open program (Lippiello and Fernandes 1986 through adjustment on film; The people such as Davies, 1999) combination to nAChR is tested in standard method.In brief, film is reconstructed in freezing storing solution, to test in buffer reagents (PBS) incubation 2 hours under competing compound (0.001nM to 100 μM) and radioligand existence in 150 μ l on ice.[ 3h]-Nicotine (L-(-)-[N-methyl- 3h]-Nicotine, 69.5Ci/mmol, Perkin-Elmer Life Sciences, Waltham, MA) for mankind α 4 β 2 binding.[ 3h]-epibatidine (52Ci/mmol, Perkin-Elmer Life Sciences) is for the binding at other nicotinic receptor subtypes place.L-[benzilic acid-4,4- 3h] quinuclidinyl dibenzene-hydroxy-acetic acid ester ([ 3h] QNB) study for muscarinic receptors bind.Incubation is stopped, to reduce non-specific binding at the upper fast filtering of manifold tissue harvester (Brandel, Gaithersburg, MD) with the GF/B strainer be dipped in advance in 0.33% polymine (w/v).The ice-cold PBS of strainer washs 3 times, is measured the radioactivity retained by liquid scintillation counting(LSC).
In conjunction with data analysis.Be expressed as per-cent overhead control in conjunction with data to combine.The repeating of each point is averaged, and maps to the log of drug level.Returned by nonlinear least-square and determine IC with GraphPadPrism software (GraphPAD, San Diego, CA) 50(produce 50% and combine the compound concentration suppressed).Ki Cheng-Prusoff formula (Cheng and Prusoff, 1973) calculates.
Calcium current leads to function test
Before each experiment 24 to 48 hours, by cell bed board in the 96 black wall dianegatives (Corning, Corning, NY) in hole, 60-100,000 cells/well.In experiment day, gently remove growth medium, buffer reagent (20mM HEPES, 7mM TRIS alkali, 4mM CaCl will be tested 2, 5mM D-Glucose, 0.8mM MgSO 4, 5mM KCl, 0.8mM MgCl 2, 120mM N-methyl D-Glucose amine, 20mM NaCl, pH 7.4, for SH-EP1-mankind α 4 β 2 cell; Or 10mM HEPES, 2.5mM CaCl 2, 5.6mM D-Glucose, 0.8mM MgSO 45.3mM KCl, 138mM NaCl, pH 7.4, containing TRIS-alkali, for whole other clone) in 200 μ L 1X FLIPR calcium 4 test agent (MolecularDevices, Sunnyvale, CA) each hole is added, at 37 DEG C of incubation plates 1 hour (being then 29 DEG C for SH-EP1-mankind α 4 β 2 cell 29 DEG C-process).In order to suppress research, add competing compound (10pM-10 μM) in the time that dyestuff adds.Remove plate from incubator, be allowed to equilibrate to room temperature.Transfer blade to FLIPR Tetra fluorometric imaging plate reader (Molecular Devices), for adding compound and monitoring fluorescence (excite 485nm, launch 525nm).Calcium current flux and positive (Nicotine) and negative control (buffer reagent is independent) are contrasted.Positive control is defined as 100% response, and the results expression of each test compound is the per-cent based on positive control.In order to suppress research, agonist Nicotine is used for SH-EP1-mankind α 4 β 2 and SH-EP1-mankind α 4 β 4 cell with 1 μM of concentration, is used for PC12 and SH-SY5Y cell with 10 μMs of concentration, and is used for TE671/RD cell with 100 μMs of concentration.
Neurotransmitter regulator
Dopamine D_2 receptors research is carried out with the line shape synaptosome deriving from rat brain describing people such as (, 1998) Bencherif in advance.Ice-cold containing the 5mM HEPES (pH7.4) of 0.32M sucrose (8mL) in collect with glass/glass homogenizer and homogenize from the line shape tissue of 2 rats (female, Sprague-Dawley, weight 150-250g).Then, in the centrifugal tissue of 1,000x g 10 minutes.Discard granule, supernatant liquor centrifugal 20 minutes in 12,500xg.By gained granule settling flux in ice-cold oxidase inhibitor (128mM NaCl, the 1.2mM KH containing perfusion buffer reagent 2pO 4, 2.4mM KCl, 3.2mM CaCl 2, 1.2mM MgSO 4, 25mMHEPES, 1mM vitamins C, 0.02mM Pargyline HCl and 10mM glucose, pH 7.4) in, in 23,000x g centrifugal 15 minutes.By final granule settling flux in pouring into buffer reagent (2mL) for instant use.
In the jolting incubator of 37 DEG C, incubation synaptosome suspension 10 minutes is to recover metabolic activity.Add [ 3h] Dopamine HCL ([ 3h] DA, activity specific=28.0Ci/mmol, NEN studies product), ultimate density 0.1 μM, another 10 minutes of 37 DEG C of incubation suspension.The aliquots containig of pouring into buffer reagent (100 μ L) and tissue (100 μ L) is loaded in the super fusion room of Brandel Suprafusion system (series 2500, Gaithersburg, MD).To pour into during buffer reagent (room temperature) pumping enters the room with the speed of about 0.6mL/min, wash 8 minutes sections.In perfusion stream, use competing compound (10pM-100nM) continue 8 minutes.Then administering nicotine (10 μMs) 48 seconds in perfusion stream.Fraction (each 12 seconds) is collected continuously from each room, to catch the peak release of basis release and agonist-induction, to redefine the baseline after agonist is used in whole experimentation.Perfusate is directly collected in scintillation vial, adds scintillation fluid to it.Scintillation counting quantitatively discharge [ 3h] DA.For each room, by fraction peak area to its baseline criteria.
Release is expressed as the per-cent of the release obtained with the contrast Nicotine that there is not rival.In each test, each test compound concentration 2 rooms are repeated; Be averaged repeating.Definition causes the compound concentration (IC of the maximum suppression of specific ion circulation half 50).
Patch clamp electrophysiology
Cell process. after incubator removing GH4C1-rat T6 ' S α 7 cell, Aspirate culture media, cell trypsinized 3 minutes, it removes from plate by gentle abrasion, wash twice with record substratum, settling flux is in 2ml external solution (composition vide infra).Cell is placed on the Dynaflow chip that is arranged on and is inverted on Zeiss microscope (Carl Zeiss Inc., Thornwood, NY) platform.Fifty-fifty, determine that whole-cell recording technique configuration needs 5 minutes.In order to avoid changing cell condition, every single load record individual cells.In order to bring out of short duration response, with Dynaflow system (Cellectricon, Inc., Gaithersburg, MD) administered compound 0.5 second, wherein each passage sends the solution of pressure-driving with 50 or 150psi.
Pincers electrophysiology. use conventional whole-cell electric current record.By glass microelectrode (5-10M Ω impedance) to form deadend (>1G Ω) at cell surface until use the whole-cell recording technique aspirated to be converted into routine.Then, vise cell at maintenance electromotive force-60mV voltage, measure the ion(ic)current that response part is used.With ADC plate 1440 (Molecular Devices), sample by the full cell currents of Axon 700A amplifier record in 5kHz in 1kHz filtering.Full cells contacting impedance is less than 20M Ω.The data gathering Clampex 10 (MolecularDevices, Sunnyvale, CA) of full cell currents completes, and result Prism 5.0 (GraphPad Software Inc., San Diego, CA) maps.Experimental data is expressed as mean value ± S.E.M., with the significance,statistical of the difference of Student t and Two Way ANOVA check analysis different condition.All experiments at room temperature (22 ± 1 DEG C) carries out.Concentration-response feature Hill formula fitting, analyzes with Prism 5.0.
Solution and medicament administration.Standard external solution contains: 120mM NaCl, 3mM KCl, 2mM MgCl 2, 2mM CaCl 2, 25mM D-Glucose, and 10mM HEPES, be adjusted to pH 7.4 with TRIS alkali.The internal solution of whole-cell recording technique is made up of following: 110mMTris bis-alkali formula phosphoric acid, 28mM TRIS alkali, 11mM EGTA, 2mM MgCl 2, 0.1mMCaCl 2, and 4mM Mg-ATP, pH 7.3.(people such as Liu, 2008).In order to cause the response of full cell currents, send compound by being moved around from the contrast solution solution moved to containing agonist by cell, solution is exchanged occur in ~ 50ms in (the peak current rise time based on 10-90%).Interval (0.5-1 minute) between special adjustment compound administration is to guarantee the stability (can not reduce function) of acceptor responsiveness, and the selection for the pipette solution in great majority research herein is also for identical object.(-)-Nicotine and vagusstoff (ACh) are purchased from Sigma-Aldrich (St.Louis, MO).Preparation every day of whole medicine is from stock solution.
In order to determine the suppression of the compounds of this invention to the electric current that ACh induces, we establish stable baseline record, use 70 μMs of ACh (normally stable 5-10 time coherent use).Then, ACh (70 μMs) is co-administered with the test compound of concentration range 1nM to 10 μM.Because the most deep change occurs tail current (ACh uses the last 0.5 second electric current measured), suppress and recover the amplitude that mapping represents tail current.
Tabulation summary
As shown in table 1, the representational compound of the present invention generally shows the suppression constant (Ki value) of 1-100mM scope for mankind α 4 β 2 and ganglionic receptor hypotype, this points out the low-affinity of the ortho position resistance binding site (being also the binding site of competitive agonist) for these receptor subtypes.But the data of table 1 also illustrate that the representational compound of the present invention suppresses the ion of these receptor subtypes to circulate effectively, its typical IC 50value is less than about 2mM and typical I maxvalue is >95%.Be summed up, the representational compound of this data display the present invention suppresses to be circulated by the ion of these receptor subtype mediates by the mechanism of the combination not being involved in resistance site, ortho position effectively.
Observe specific pharmacological response can according to and depend on selected particular active compounds or whether there is pharmaceutical carrier, and preparaton type used and mode of administration and change, it is desired that the results change of described expectation or difference are that the present invention puts into practice.
Although particular implementation of the present invention has been described in detail in the description, the present invention has been not limited to this.Above-mentioned detailed description provides as example of the present invention, and should not be construed as formation to any restriction of the present invention.Those skilled in the art will know its various modification, within the whole modification not deviating from purport of the present invention all expect to be included in the scope of claims.

Claims (15)

1. formula I:
Formula I
Wherein
R 1and R 2h, C individually separately 1-6alkyl, or the C of aryl-replacement 1-6alkyl, or R 1and R 2the nitrogen-atoms be connected to them is combined to form 3 to 8 rings, and described ring optionally can use following replacement: C 1-6alkyl, aryl, C 1-6alkoxyl group, or aryloxy substituting group;
R 3h, C 1-6alkyl, or C 1-6the C of alkoxyl group-replacement 1-6alkyl;
R 4, R 5, R 6and R 7h, C individually separately 1-6alkyl, or C 1-6alkoxyl group;
L 1be selected from following linker kind: CR 8r 9, CR 8r 9cR 10r 11, and O;
L 2be selected from following linker kind: CH 2, CH 2cH 2, CH 2cH 2cH 2, and CH 2cH 2cH 2cH 2;
R 8, R 9, R 10and R 11hydrogen or C individually separately 1-6alkyl; With
Dotted line points out optional double bond;
Or its pharmacy acceptable salt.
2. the compound of claim 1, wherein R 1be H and R 2c 1-6alkyl.
3. the compound of claim 1 or 2, wherein R 3c 1-6alkyl.
4. the compound any one of claim 1-3, wherein R 4, R 5, R 6and R 7each H naturally.
5. the compound any one of claim 1-4, wherein L 1cR 8r 9, and R 8and R 9each hydrogen naturally.
6. the compound any one of claim 1-5, wherein L 2cH 2cH 2.
7. the compound any one of claim 1-6, wherein said dotted line is singly-bound.
8. pharmaceutical composition, comprises compound claimed any one of claim 1-7 and pharmaceutically acceptable carrier.
9. treat or prevent the method for disease or the illness mediated by neuronic nAChR, comprise compound claimed any one of administration claim 1-7.
10. the method for claim 9, wherein said disease or illness are IBS-D, OAB, nicotine addiction, and smoking stops, depressed, major depressive disorder, or hypertension.
11. compound claimed any one of claim 1-7 is for the preparation of the purposes of medicine, described medicine is used for the treatment of or prevents the disease that mediated by neuronic nAChR or illness.
12. the purposes of claim 11, wherein said disease or illness are IBS-D, OAB, nicotine addiction, and smoking stops, depressed, major depressive disorder, or hypertension.
Compound claimed any one of 13. claim 1-7, as active therapeutic agent.
14. compound claimed any one of claim 1-7, in the treatment of the disease that mediated by neuronic nAChR or illness or prevention.
The compound of 15. claims 13 or 14, wherein said disease or illness are IBS-D, OAB, nicotine addiction, and smoking stops, depressed, major depressive disorder, or hypertension.
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